Chemico-Biological Interactions 189 (2011) 100–106

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Chemico-Biological Interactions
journal homepage: www.elsevier.com/locate/chembioint

Ameliorative potential of S-allyl cysteine on oxidative stress

in STZ induced diabetic rats
Ganapathy Saravanan a,b,∗ , Ponnusamy Ponmurugan a,c
a
Research and Development Centre, Bharathiyar University, Coimbatore, Tamil Nadu 641046, India
b
Department of Biochemistry, Centre for Biological Science, K.S.R. College of Arts and Science, Thokkavadi, Tiruchengode, Tamil Nadu 637215, India
c
Department of Biotechnology, K.S.R. College of Technology, Thokkavadi, Tiruchengode, Tamil Nadu 637215, India

a r t i c l e i n f o a b s t r a c t

Article history: Increased oxidative stress and impaired antioxidant defense mechanism are important factors in the
Received 24 July 2010 pathogenesis and progression of diabetes mellitus and other oxidant-related diseases. The present study
Received in revised form 5 October 2010 was undertaken to evaluate the possible protective effects of S-allyl cysteine (SAC) against oxidative
Accepted 7 October 2010
stress in streptozotocin (STZ) induced diabetic rats. SAC was administered orally for 45 days to control
Available online 14 October 2010
and STZ induced diabetic rats. The effects of SAC on glucose, plasma insulin, thiobarbituric acid reactive
substances (TBARS), hydroperoxide, superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase
Keywords:
(GPx), reduced glutathione (GSH), oxidized glutathione (GSSG) and GSH/GSSG ratio were studied. The lev-
Antioxidants
Diabetes mellitus
els of glucose, TBARS, hydroperoxide, and GSSG were increased significantly whereas the levels of plasma
Streptozotocin insulin, reduced glutathione, GSH/GSSG ratio, superoxide dismutase, catalase and GPx were decreased
S-allyl cysteine in STZ induced diabetic rats. Administration of SAC to diabetic rats showed a decrease in plasma glucose,
TBARS, hydroperoxide and GSSG. In addition, the levels of plasma insulin, superoxide dismutase, cata-
lase, GPx and reduced glutathione (GSH) were increased in SAC treated diabetic rats. The above findings
were supported by histological observations of the liver and kidney. The antioxidant effect of SAC was
compared with glyclazide, a well-known antioxidant and antihyperglycemic drug. The present study
indicates that the SAC possesses a significant favorable effect on antioxidant defense system in addition
to its antidiabetic effect.
© 2010 Elsevier Ireland Ltd. All rights reserved.

1. Introduction
Diabetes mellitus (DM) is the most significant chronic disease
and cause of death in modern society. Diabetes mellitus comprises
a group of chronic disorders characterized by hyperglycemia or
diminished insulin secretion, or both. DM involves high level of
blood glucose, which contributes to an increase in free radical
production [1]. DM is a prevalent systemic disease affecting a sig-
nificant proportion of the population worldwide. The World Health
Organization (WHO) predicts that by the year 2025, 300 million
people will have diabetes mellitus [2]. Studies conducted in India
in the last decade have highlighted that not only in the prevalence
of diabetes high but also it is increasing rapidly in the urban pop-
ulation [3]. The International Diabetes Federation (IDF) estimates
the total number of diabetic subjects to be around 40.9 million in
Abbreviations: SAC, S-allyl cysteine; STZ, streptozotocin; kg, kilogram.
∗ Corresponding author at: Department of Biochemistry, Centre for Biological
Science, K.S.R. College of Arts and Science, Thokkavadi, Tiruchengode, Tamil Nadu
637215, India. Tel.: +91 9843954422.
E-mail address: saravana bioc@rediffmail.com (G. Saravanan).
India and this is further set to rise to 69.9 million by the year 2025
[4].
Oxidative stress can be associated to an increased rate of reactive
oxygen species (ROS) generation, a decrease of antioxidant defense
or a combination of both. ROS-mediated alterations include damage
to cells, tissues or organs and are proposed as the major factors
in the mechanism of several diseases [1]. An increased production
of oxygen-derived free radicals and the decrease in the activity of
free radical scavenger system have been reported in diabetes [5].
It has also been proposed that an increase in oxidative stress could
contribute to tissue damage in diabetes [6]. Over the past decade
there has been an increased interest in oxidative stress and its role
in the development of complications of diabetes. Apart from the
traditional antidiabetic treatment, antioxidant therapy may benefit
in diabetes.
In recent years, considerable focus has been given to an intensive
search for novel type of antioxidants present in plants for treating
disease [7,8] and some of the plants are used by the population
as anti diabetic remedies [9]. Management of diabetes without
any side effects is still a challenge to the medical system. There
is an increasing demand by patients to use the natural products
with antidiabetic activity, because insulin and oral hypoglycemic

0009-2797/$ – see front matter © 2010 Elsevier Ireland Ltd. All rights reserved.
doi:10.1016/j.cbi.2010.10.001
 G. Saravanan, P. Ponmurugan / Chemico-Biological Interactions 189 (2011) 100–106
drugs possess undesirable side effects [10]. Plants and its active
compounds provide useful sources for the development of drugs
in the treatment of diabetes mellitus. Clinical research has con-
firmed the efficacy of several plants and its active compounds in the
modulation of oxidative stress associated with diabetes mellitus
[11].
S-allyl cysteine (SAC), a sulphur containing amino acid, derived
from garlic (Allium sativum Linn, Liliaceae), may constitute an alter-
native to insulin since both long- and short-term treatments with
this compound correct the hyperglycemia that occurs in diabetic
model [12]. Treatment with SAC also prevents the occurrence of
various complications of diabetes in rats by changing the blood
glucose and protein metabolism [12]. SAC normalizes hepatic car-
bohydrate metabolism [13] and plasma and tissue glycoprotein
level [14] in diabetic rats. However, no studies have specifically
addressed the efficacy of SAC in STZ-induced oxidative stress in rats.
Thus, the present manuscript addresses the protective effect of SAC
on lipid peroxidation, antioxidant defense system and histopatho-
logical changes in tissues of STZ-induced diabetic rats. The effects
produced by these treatments are compared with standard drug
glyclazide.
2. Materials and methods
2.1. Animals
Male Wistar rats of body weight 150–180 g were used for this
study. The animals were maintained in the animal house, Sas-
tra University, Thanjavore, India and fed on standard pellet diet
(Amrut, Pune, India) and water ad libitum. The protocol of this
study was approved by Institutional Ethical Committee of Sastra
University.
2.2. Chemicals
SAC (99%) was purchased from LGC Prochem, Bangalore, India.
Streptozotocin was purchased from Himedia, Bangalore, India. All
other chemicals used were of analytical grade.
2.3. Induction of diabetes
Diabetes was induced [15] in overnight fasted adult Wistar
strain albino male rats weighing 150–180 g by a single intraperi-
toneal injection of 55 mg/kg streptozotocin. Streptozotocin was
dissolved in 0.1 M citrate buffer (pH 4.5). Hyperglycemia was con-
firmed by the elevated glucose levels (above 250 mg/dl) in blood,
determined at 72 h and then on day 7 after injection.
2.4. Experimental design
The rats were divided into four groups each comprising a mini-
mum of six rats.
Group 1: normal control rats;
Group 2: diabetic control rats;
Group 3: STZ-treated rats given SAC (150 mg/kg body weight)
orally using an intragastric tube daily for a period of 45 days [12]
and
Group 4: STZ-treated rats given glyclazide (5 mg/kg b.w. of rat)
orally using an intragastric tube daily for a period of 45 days [16].
After the last treatment (45 days), rats were fasted overnight and
sacrificed by cervical decapitation. Blood was collected and plasma
was obtained after centrifugation. It was used for the estimation of
glucose. Liver and kidney tissues were excised immediately from
101
the rats and stored in ice-cold containers. They were then homog-
enized with buffer, centrifuged and the supernatant was collected.
Biochemical estimations were carried out in the homogenates.
2.5. Biochemical estimations
2.5.1. Determination of blood glucose and plasma insulin
Plasma glucose was estimated colorimetrically by using com-
mercial diagnostic kits (Sigma Diagnostics (I) Pvt. Ltd., Baroda,
India). Plasma insulin assay was assayed by enzyme linked
immunosorbent assay kit (ELISA, Boerhriger Mannheim, Germany).
2.5.2. Determination of lipid peroxidation
The level of thiobarbituric acid reactive substances (TBARS)
was estimated by the method of Fraga et al. [17]. In brief, 0.5 ml
of homogenate was treated with 2 ml (1:1:1 ratio) TBA–TCA–HCl
reagent (thiobarbituric acid, 0.37%, 0.25 N HCl, 15% TCA) placed for
15 min in a water bath and cooled. The absorbance of the clear
supernatant was measured against reference blank at 535 nm. Val-
ues were expressed as mM/100 g-tissue.
The hydroperoxide was determined by the method of Jiang
et al. [18]. Tissue homogenate (0.1 ml) was treated with 0.9 ml of
Fox reagent (88 mg of butylated hydroxy toluene (BHT), 7.6 mg of
xylenol orange and 0.8 mg of ammonium iron sulphate were added
to 90 ml of methanol and 10 ml of 250 mM sulphuric acid) and incu-
bated at 37 ◦ C for 30 min. Then the absorbance was read at 560 nm.
Hydroperoxides were expressed as mM/100 g-tissue.
2.5.3. Determination of reduced glutathione
Reduced glutathione was measured according to the method of
Beutler and Kelley [19]. The technique involved in protein precip-
itation by metaphosphoric acid and spectrophotometric assay at
412 nm of the yellow derivative obtained by the reaction of super-
natant with 5, 5 -dithio-bis-2-nitrobenzoic acid (DTNB).
2.5.4. Determination of oxidized glutathione
Oxidized glutathione was measured according to the method
described by Aseni et al. [20] based on the principle of glutathione
reductase enzyme reducing GSSG to GSH with the concomitant
oxidation of NADPH to NADP+ . To 0.9 ml of 1.75 mol/l K3 PO4
buffer (pH 7.0) containing 20 mmol/l NEM was added 0.05 ml of
sample extract and 0.025 ml of 10 mg/ml of NADPH–Na solution.
Absorbance at 340 nm was measured for 30 s immediately after
addition of 0.025 ml of (10 mg/ml) glutathione reductase (GR) to
the assay mixture.
2.5.5. Determination of superoxide dismutase
Superoxide dismutase (SOD) activity was assayed by the method
of Kakkar et al. [21]. 0.5 ml of tissue homogenate was diluted
with 1 ml of water. In this mixture, 2.5 ml of ethanol and 1.5 ml
of chloroform (all reagents chilled) were added and shaken for
1 min at 4 ◦ C then centrifuged. The enzyme activity in the super-
natant was determined. The assay mixture contained 1.2 ml of
sodium pyrophosphate buffer (0.025 M, pH 8.3), 0.1 ml of 186 ␮m
phenazine methosulphate (PMS), 0.3 ml of 30 ␮m nitroblue tetra-
zolium salt (NBT), 0.2 ml of 780 ␮m NADH, appropriately diluted
enzyme preparation and water in a total volume of 3 ml. Reaction
was started by the addition of NADH. After incubation at 30 ◦ C for
90 s the reaction was stopped by the addition of 1 ml glacial acetic
acid. The reaction mixture was stirred vigorously and shaken with
4 ml of n-butanol. The intensity of the chromogen in the butanol
layer was measured at 560 nm against butanol blank. A system
devoid of enzyme served as control. One unit of the enzyme activ-
ity is defined as the enzyme reaction, which gave 50% inhibition of
NBT reduction in 1 min under the assay condition.
102 G. Saravanan, P. Ponmurugan / Chemico-Biological Interactions 189 (2011) 100–106

Table 1
Levels of lipid peroxides and hydroperoxides in liver of control and experimental
groups of rats.

Groups TBARS Hydroperoxides

(mM/100 g) of (mM/100 g of
tissue tissue)

Control 8.84 ± 1.05 2.38 ± 0.84

Diabetic 14.26 ± 1.53a , * 4.32 ± 1.14a , *
Diabetic + SAC 9.93 ± 0.86b * 2.16 ± 0.95b , *
Diabetic + glyclazide 8.98 ± 1.3c * 2.47 ± 1.32c , *

Values are mean ± SD, n = 6.

a
Significantly different from control.
b
Significantly different from diabetic control.
c
Significantly different from diabetic control.
*
P < 0.001.
Fig. 1. Levels of blood glucose in control and experimental groups of rats. Values
are mean ± SEM, n = 6. a Significantly different from control. b Significantly different
Table 2
from diabetic control. c Significantly different from diabetic control. *P < 0.05.
Levels of lipid peroxides and hydroperoxides in kidneys of control and experimental
groups of rats.
2.5.6. Determination of catalase Groups TBARS Hydroperoxides
Catalase level was determined by Aebi’s modified colorimetric (mM/100 g of (mM/100 g of
method [22]. The reaction mixture (1.5 ml, vol.) contained 1.0 ml tissue tissue)
of 0.01 M phosphate buffer (pH 7.0), 0.1 ml of tissue homogenate Control 10.87 ± 1.53 2.83 ± 1.21
and 0.4 ml of 2 M H2 O2 . The reaction was stopped by the addition of a
Diabetic 16.09 ± 1.47a , *** 4.71 ± 0.77a , **
2.0 ml of dichromate–acetic acid reagent (5% potassium dichromate b
Diabetic + SAC 9.69 ± 0.73b , *** 3.05 ± 0.72b , *
b
Diabetic + glyclazide 10.33 ± 1.35c , *** 2.52 ± 0.75c , **
and glacial acetic acid were mixed in 1:3 ratio). Then the absorbance
was read at 620 nm; CAT activity was expressed as ␮M of H2 O2 Values are mean ± SD, n = 6.
a
consumed/min/mg protein. Significantly different from control.
b
Significantly different from diabetic control.
c
Significantly different from diabetic control.
2.5.7. Determination of glutathione peroxidase *
P < 0.05.
**
GPx activity was measured by Paglia and Valentine’s method P < 0.01.
***
[23] The reaction mixture contained 2.6 ml of 100 mmol/l phos- P < 0.001.

phate buffer (pH 7.0) with 3 mmol/l EDTA, 0.05 ml of 10 mg/ml GSH
solution, 0.1 ml GR (10 mg/ml), 0.05 ml (10 mg/ml) NADPH–Na salt, Fig. 3 summarizes the concentration of GSSG and the level
0.1 ml 90 mmol/l hydrogen peroxide solution and 0.1 ml of sample. of GSH/GSSG ratio in control and experimental rats. There was
The GPx activity was monitored by the decrease in absorbance due a significant (P < 0.05) increased level of GSSG and concomitant
to the consumption of NADPH. The absorbance was read at 340 nm. decreased in the level of GSH/GSSG in diabetic rats when compared
to control rats. Oral treatment of SAC and glyclazide tended to bring
2.6. Statistical analysis GSSG and GSH/GSSG ratio towards near normal levels.
Tables 3 and 4 summarize the activities of superoxide dismutase
All the results in tables were expressed as the mean ± SD for (SOD), catalase (CAT) and glutathione peroxidase (GPx) in the liver
six animals in each group and figure results were expressed in and kidneys of control and experimental groups of rats. The activ-
mean ± SEM. All the grouped data were statistically evaluated with ities of SOD, CAT and GPx in liver and kidneys were significantly
SPSS\10.0 software. Hypothesis testing methods included one way (P < 0.05) lower in diabetic control rats compared to diabetic rats.
analysis of variance (ANOVA) followed by least significant differ- Treatment with SAC and glyclazide showed a significant increase
ence (LSD) test; significance levels at P < 0.01, 0.05 and 0.001 were in activities of SOD, CAT and GPx in liver and kidneys of diabetic
considered to indicate statistical significance. rats.
Histopathological studies revealed normal central vein and radi-
ating hepatocytes in control liver (Fig. 4A) whereas diabetic control
3. Results
rats showed vascular congestion and mononuclear cellular infil-
tration (Fig. 4B). SAC and glyclazide treated rats revealed normal
Fig. 1 shows the levels of glucose and plasma insulin in control
and experimental animals. There was a significant (P < 0.05) eleva-
tion in blood glucose level with a significant decrease in plasma
insulin levels in STZ induced diabetic rats, compared with control
rats. Administration of SAC and glyclazide tended to bring blood
glucose and plasma insulin towards near normal levels.
Tables 1 and 2 show the concentration of TBARS, hydroperox-
ides in liver and kidneys of control and experimental groups of rats.
The levels of TBARS and hydroperoxides in diabetic rats were sig-
nificantly (P < 0.05) higher than control rats, whereas diabetic rats
treated with SAC and glyclazide restored the altered values to the
near normal.
Fig. 2 shows the levels of reduced glutathione in liver and kid-
neys of control and experimental groups of rats. The decreased
concentration (P < 0.05) of GSH was observed in diabetic rats.
Fig. 2. Levels of GSH in liver and kidneys of control and experimental groups of rats.
Administration of SAC and glyclazide tends to bring the GSH level Values are mean ± SEM, n = 6. a Significantly different from control. b Significantly dif-
to near normal. ferent from diabetic control. c Significantly different from diabetic control. *P < 0.05.
 G. Saravanan, P. Ponmurugan / Chemico-Biological Interactions 189 (2011) 100–106 103

Table 3
Activities of superoxide dismutase, catalase and glutathione peroxidase in the liver
of control and experimental groups of rats.

Groups SOD CAT GPx

Control 7.78 ± 1.06 13.4 ± 0.72 7.10 ± 0.82

Diabetic 4.88 ± 1.08a , ** 9.33 ± 1.20a , *** 3.79 ± 1.17a , ***
Diabetic + SAC 6.82 ± 0.38b , * 11.37 ± 0.93b , * 5.54 ± 0.94b , *
Diabetic + glyclazide 7.16 ± 1.57c , * 12.28 ± 1.67c , ** 5.95 ± 0.99c , **

Values are mean ± SD, n = 6.

Activity is expressed as: 50% of inhibition of epinephrine auto oxidation per min
for SOD; ␮mole of hydrogen peroxide decomposed per min per mg of protein for
catalase; ␮mole of glutathione oxidized per min per mg of protein for GPx.
a
Significantly different from control. Fig. 3. Level of GSSG and GSH/GSSG ratio in control and experimental rats. Values
b
Significantly different from diabetic control. are mean ± SEM, n = 6. a Significantly different from control. b Significantly different
c
Significantly different from diabetic control. from diabetic control. c Significantly different from diabetic control. *P < 0.05.
*
P < 0.05.
**
P < 0.01.
***
P < 0.001. architecture with regenerating hepatocytes (Fig. 4C and D). Kid-
ney of control rat showed normal tubules and glomerulus (Fig. 5A),
Table 4 whereas kidney of diabetic control rat revealed intertubular haem-
Activities of superoxide dismutase, catalase and glutathione peroxidase in kidneys orrhage and mononuclear cellular infiltration (Fig. 5B). Kidney
of control and experimental groups of rats. of SAC and glyclazide treated rats revealed normal tubules and
Groups SOD CAT GPx glomerulus (Fig. 5C and D).
Control 10.15 ± 1.61 9.76 ± 0.55 7.54 ± 0.86
Diabetic 5.97 ± 1.47a , *** 4.38 ± 1.46a , *** 3.18 ± 0.90a , *** 4. Discussion
Diabetic + SAC 8.76 ± 0.96b , * 7.98 ± 0.83b , *** 5.56 ± 1.77b , *
Diabetic + glyclazide 9.24 ± 1.94c , ** 8.26 ± 2.01c , *** 6.04 ± 0.77c , ** Streptozotocin (STZ)-induced hyperglycemia in animals is con-
Values are mean ± SD, n = 6. sidered to be a good model for the preliminary screening of agents
Activity is expressed as: 50% of inhibition of epinephrine auto oxidation per min active against diabetes [24]. In experiment with many animal
for SOD; ␮mole of hydrogen peroxide decomposed per min per mg of protein for
species [25], streptozotocin produces permanent diabetes that
catalase; ␮mole of glutathione oxidized per min per mg of protein for GPx.
a impersonates the pathological status found in human diabetes [26].
Significantly different from control.
b
Significantly different from diabetic control. In the present study, the intraperitoneal administration of strepto-
c
Significantly different from diabetic control. zotocin effectively induced diabetes mellitus in rats. STZ-induced
*
P < 0.05. experimental diabetes may exhibit most of the diabetic compli-
**
P < 0.01.
***
cations mediated through oxidative stress [27]. During diabetic
P < 0.001.
state, increased generation of ROS occur and cause an imbalance

Fig. 4. (A) Liver section of normal control showing normal central vein and radiating hepatocytes. H&E 400×. (B) Liver section of diabetic rats showing vascular congestion and
mononuclear cellular infiltration (arrow mark shows mononuclear cellular infiltration). H&E 400×. (C) Liver section from SAC treated animals revealing normal architecture
with regenerating hepatocytes. H&E 100×. (D) Liver section from glyclazide treated animals revealing normal architecture with regenerating hepatocytes. H&E 400×.
104 G. Saravanan, P. Ponmurugan / Chemico-Biological Interactions 189 (2011) 100–106

Fig. 5. (A) Kidney section from control showing normal tubules and glomerulus. H&E 400×. (B) Kidney section diabetic control revealing intertubular haemorrhage and
mononuclear cellular infiltration (arrow mark shows intertubular haemorrhage). H&E 400×. (C) Kidney section SAC treated animals revealing normal tubules and glomerulus.
H&E 400×. (D) Kidney section glyclazide treated animals revealing normal tubules and glomerulus. H&E 400×.

between the oxidant and antioxidant status [28]. Administration
of STZ resulted in a significant increase in the blood glucose level
and reduction in plasma insulin level. Sustained hyperglycemia has
been identified as a principle mediator of increased reactive oxy-
gen species generation in diabetes [29]. Oral administration of SAC
decreased the blood glucose level and increased plasma insulin
level in diabetic rats. This may be due to the insulin like effect of
SAC on peripheral tissues, either by promoting glucose uptake and
metabolism, or by inhibiting hepatic gluconeogenesis [12].
Increased free radical observed in diabetic rats is attributed to
chronic hyperglycemia that damage antioxidant defense system
[30]. Free radicals may also be formed via the auto-oxidation of
unsaturated lipids in plasma and membrane lipids. The free rad-
ical produced may react with polyunsaturated fatty acids in cell
membrane leading to lipid peroxidation. Lipid peroxidation will in
turn result in elevated production of free radicals [31]. Increased
lipid peroxidation impairs membrane functions by decreasing
membrane fluidity and changing the activity of membrane-bound
enzymes and receptors [32]. Its products are harmful to most of
the cells in the body and are associated with a variety of diseases
[33]. Our present study showed a significant increase of tissue thio-
barbituric acid reactive substances (TBARS) and hydroperoxides
(HP) level in diabetic rats. The increased TBARS content of dia-
betic rats suggests that peroxidative injury may be involved in the
development of diabetic complications. TBARS and hydroperoxides
levels in liver and kidneys were significantly lower in the SAC and
glyclazide-treated group compared to the diabetic control rats. The
above result suggests that the SAC may exert antioxidant activities
and protect the tissues from lipid peroxidation.
GSH is a major endogenous antioxidant which counterbalance
free radical mediated damage. Studies have shown that the tissue
GSH concentrations of STZ-induced diabetic rats are significantly
lower when compared with the control rats [34]. It is well known
that GSH is involved in the protection of normal cell structure and
function by maintaining the redox homeostasis, quenching of free
radicals and participating in detoxification reactions. It is a direct
scavenger of free radicals as well as a co-substrate for peroxide
detoxification by glutathione peroxidases [35]. Loven et al. [36]
had suggested that the decrease in tissue GSH could be the result
of decreased synthesis or increased degradation of GSH by oxida-
tive stress in diabetes. Increased oxidative stress, resulting from
a significant increase in aldehydic products of lipid peroxidation
has probably decreased the tissue GSH content [35]. In the present
study, the elevation of GSH levels in liver and kidneys was observed
in the SAC and glyclazide treated diabetic rats. This indicates that
the SAC and glyclazide can either increase the biosynthesis of GSH
or reduce the oxidative stress leading to less degradation of GSH,
or have both effects.
GSH and GSSG levels are commonly used markers for oxida-
tive stress. The decrease in GSH content, which is shown in the
results of the present work, seems to be due to its oxidation to
GSSG (oxidized glutathione) [37]. GSSG appears to be released from
most cells as a consequence of oxidative stress so that an oxida-
tion of the cellular pool could shift the balance of GSH and GSSG
efflux and change the extra cellular redox state. This indicates that
GSH/GSSG redox system plays a significant role in diabetes. In other
words, a low GSH/GSSG ratio suggests increased oxidative stress.
The GSH/GSSG ratio seems to play a major role in the modulations of
glucose homeostasis in diabetes [38]. GSH infusion has been shown
to increase GSH/GSSG ratio and potentiate the insulin action in
diabetic patients [33]. Thus, any drug that replenishes glutathione
may be able to reverse the oxidative damage caused in diabetes
mellitus and prevent the associated disorders. SAC, a known antiox-
idant probably strengthened the antioxidant status as evidenced by
increase in the GSH levels.
In diabetic state, insulin deficiency caused the impairment of
glucose utilisation leading to an increased generation of oxygen
free radicals [35]. Studies have reported on the reduction of tissue
SOD activities in STZ-induced diabetic rats [39]. SOD is an important
defense enzyme which catalyses the dismutation of superoxide
 G. Saravanan, P. Ponmurugan / Chemico-Biological Interactions 189 (2011) 100–106
radicals to produce H2 O2 and molecular oxygen [40], hence dimin-
ishing the toxic effects caused by their radical. Wohaieb et al. [41]
had suggested that the reactive oxygen free radicals could inacti-
vate and reduce the tissue SOD activities. The observed decrease
in SOD activity could result from inactivation by H2 O2 or by gly-
cation of enzymes [42]. Oral treatment of SAC caused a significant
increase in SOD activities in diabetic rats. This means that the SAC
can reduce reactive oxygen free radicals and improve the activities
of the tissue antioxidant enzymes.
Catalase (CAT) is a hemeprotein which catalyses the reduction
of hydrogen peroxides and protects the tissues from highly reac-
tive hydroxyl radicals [43]. The decrease in CAT activity could result
from inactivation by glycation of enzyme [44]. CAT reduces hydro-
gen peroxide produced by dismutation reaction. It also prevents
the generation of hydroxyl radicals thereby protecting the cellular
constituents from oxidative damage in peroxisomes. The reduced
activity of CAT in STZ-treated rats results in the accumulation of
H2 O2 , which produces deleterious effects. In the present study, it
was observed that the SAC caused a significant increase in the activ-
ity of CAT in diabetic rats. This action is predominantly due to the
antioxidant nature of SAC and could involve a mechanism related
to scavenging activity.
Glutathione peroxidase (GPx), a selenium containing enzyme
present in significant concentrations, detoxifies H2 O2 to H2 O
through the oxidation of reduced glutathione [34]. GPx works
together with glutathione in the decomposition of H2 O2 or other
organic hydroperoxides to non-toxic products at the expense of
reduced glutathione [35]. Reduced activities of GPx may result
from radical-induced inactivation and glycation of the enzyme [34].
The low activity of GPx could be directly explained by the low
content of glutathione found in diabetic state, since glutathione
is a substrate and cofactor of GPx. Reduced activities of GPx in
the liver and kidney have been observed during diabetes and this
may result in a number of deleterious effects due to the accu-
mulation of toxic products. In this context, other workers also
reported a decrease in the activity of GPx in the liver and kid-
neys of diabetic rats [45]. In the present study, administration
of SAC and glyclazide increased the activities of GPx in diabetic
rats.
In the present study, the histopathological examinations of
the liver and kidneys of diabetic rats showed vascular conges-
tion and mononuclear cellular infiltration of the hepatocytes,
as well as areas of intertubular haemorrhage and mononuclear
cellular infiltration. The reaction is provoked by the increased
production of highly reactive intermediates of STZ, which are nor-
mally detoxified by endogenous GSH [46]. The above pathological
changes were reduced in diabetic rats treated with SAC and gly-
clazide.
In conclusion, the present investigation showed that SAC pos-
sesses an antioxidant activity and also protects lipid peroxidation
and enhances its effect on cellular antioxidant defense. This activity
contributes to the protection against oxidative damage in STZ-
induced diabetes.
Conflict of interest statement
None.
Acknowledgements
The authors thank the management of K.S.R. College of Arts and
Science, Tiruchengode, India and Dr. N. Kannan, principal of this
college for their encouragement and also the management of Sastra
University, Thanjavore, India for providing facilities to do animal
studies.
105
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