Effect of chronic exposure to aspartame on oxidative stress in the brain of albino rats
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Effect of chronic exposure to aspartame on oxidative stress in the
brain of albino rats
ASHOK IYYASWAMY and SHEELADEVI RATHINASAMY*
Department of Physiology, Dr. ALM PG Institute of Basic Medical Sciences,
University of Madras, Sekkizhar campus, Taramani, Chennai 600 113, India
*Corresponding author (Fax, +91-44-24540709; Email, drsheeladevi@unom.ac.in)
This study was aimed at investigating the chronic effect of the artificial sweetener aspartame on oxidative stress in
brain regions of Wistar strain albino rats. Many controversial reports are available on the use of aspartame as it
releases methanol as one of its metabolite during metabolism. The present study proposed to investigate whether
chronic aspartame (75 mg/kg) administration could release methanol and induce oxidative stress in the rat brain. To
mimic the human methanol metabolism, methotrexate (MTX)-treated rats were included to study the aspartame
effects. Wistar strain male albino rats were administered with aspartame orally and studied along with controls and
MTX-treated controls. The blood methanol level was estimated, the animal was sacrificed and the free radical changes
were observed in brain discrete regions by assessing the scavenging enzymes, reduced glutathione, lipid peroxidation
(LPO) and protein thiol levels. It was observed that there was a significant increase in LPO levels, superoxide
dismutase (SOD) activity, GPx levels and CAT activity with a significant decrease in GSH and protein thiol.
Moreover, the increases in some of these enzymes were region specific. Chronic exposure of aspartame resulted in
detectable methanol in blood. Methanol per se and its metabolites may be responsible for the generation of oxidative
stress in brain regions.
[Iyyaswamy A and Rathinasamy S 2012 Effect of chronic exposure to aspartame on oxidative stress in the brain of albino rats. J. Biosci.
37 679–688] DOI 10.1007/s12038-012-9236-0
1. Introduction blood–brain barrier, which protects the brain from large
fluctuations in plasma aspartate (Maher and Wurtman
Aspartame (L-aspartyl-L-phenylalanine methyl ester), a low- 1987). Phenylalanine is an amino acid essential to the pro-
calorie sweetener, is about 180 times sweeter than normal duction of monoamines in the brain and is found in nearly all
sugar and is used in over 5000 foods and beverages in protein foods (Fernstrom et al. 1983). On the other hand, due
today’s market. The use of aspartame by diabetic individuals to the high levels of phenylalanine in blood, consumption of
is increasing; it is widely used in the weight loss regime, and aspartame may cause brain damage (Haschemeyer and
approximately 200 million people consume aspartame Haschemeyer 1973). Certain brain amino acid levels have
worldwide (Shapiro 1988). In 1965 aspartame was discov- been shown to be increased after aspartame consumption
ered by James Schlatter of the G.D. Searle Company (Dailey et al. 1991; Diomede et al. 1991; Yokogoshi et al.
(Garriga and Metcalfe 1988). Aspartame has a biological 1984). Although methanol is released during aspartame di-
effect even at the recommended daily dose (Gombos et al. gestion, it only accounts for 10% of the aspartame molecule
2007). Approximately 50% of the aspartame molecule is (Stegink et al. 1983), and among the metabolites, methanol
phenylalanine, 40% is aspartic acid and 10% is methanol is a toxicant that causes systemic toxicity (Kruse 1992).
(Newsome 1986). Aspartic acid is a metabolite of aspartame Various neurochemical effects due to aspartame con-
that is an excitatory amino acid and is normally found in sumption have been reported (Coulombe and Sharma 1986;
high levels in the brain. These levels are controlled by the Pan-Hou et al. 1990; Goerss et al. 2000). Their adverse
Keywords. Aspartame; blood methanol; oxidative stress; rat folate-deficient model; free radical
http://www.ias.ac.in/jbiosci J. Biosci. 37(4), September 2012, 679–688, * Indian Academy of Sciences 679
Published online: 3 August 2012
680 Ashok Iyyaswamy and Sheeladevi Rathinasamy
neurological effects include headache (Johns 1986), insom- is to investigate whether the chronic oral administration
nia and seizures after ingestion of aspartame, which are also of aspartame (75 mg/kg) can release methanol as a by-product
accompanied by the alterations in regional concentrations of after metabolism and whether it induces oxidative stress in the
catecholamine (Coulombe and Sharma 1986). Previous find- rat brain regions after aspartame administration.
ing have shown that aspartate may lead to neurotoxicity
through sustained contact with the receptors, such as gluta- 2. Materials and methods
mate producing an excitotoxic effect (Olney et al. 1980).
Large doses of both aspartame as well as these individual
2.1 Animals
metabolites have been tested in humans and other animals,
with controversial reports. It has been reported that not only
Wistar strain male albino rats (200–220 gm) were main-
the metabolites of methanol, but methanol per se as well, is
tained under standard laboratory conditions with water and
toxic to the brain (Jeganathan and Namasivayam 1998). The
food. For the folate-deficient group, folate-deficient diet was
primary metabolic fate of methanol is the direct oxidation to
provided for 45 days prior to the experiment and MTX was
formaldehyde and then into formate. The toxic effects of
administered for a week before the experiment. The animals
methanol in humans are due to the accumulation of its
were handled according to the principles of laboratory care
metabolite formate (Tephly and McMartin 1984; Tephly
framed by the committee for the purpose of Control and
1991). The severity of clinical findings in methanol intoxi-
Supervision of Experiments on Animals (CPCSEA),
cation correlated better with formate levels (Osterloh et al.
Government of India. Prior to the experimentation, proper
1986). Formate is metabolized twice as fast in rat as in approval was obtained from the Institutional Animal Ethical
monkey (McMartin et al. 1978). The metabolism of formate Committee (No: 01/032/2010).
is mediated through a tetra-hydro-folate-dependent pathway
(Eells et al. 1982). Humans and non-human primates are
uniquely sensitive to methanol poisoning because of their 2.2 Chemicals
low liver folate content (Johlin et al. 1987). There are pro-
found differences in the rate of formate oxidation in different Aspartame, methotrexate and malondialdehyde were pur-
species which determine their sensitivity to methanol chased from Sigma-Aldrich.Co., St. Louis, USA. All the
(Mcmartin et al. 1978; Eells et al. 1983). In non-human other chemicals were of analytical grade obtained from
primates and humans, alcohol dehydrogenase mediates this Sisco research laboratory, Mumbai, India.
reaction (Makar et al. 1990). In rats and other non-primate
species, this reaction is mediated by catalase. Microsomal 2.3 Experimental design
oxidizing system is reported to be responsible for free radical
generation. In addition to that, inhibition of cytochrome The rats were divided in to three groups, namely, saline
oxidase by formate leads to the generation of superoxide, control, MTX-treated control, and MTX-treated aspartame-
peroxyl and hydroxyl radicals. Tephly (1991) also proved administered groups. Each group consisted of six animals.
that a catalase system is also one of the major pathways of Aspartame mixed in sterile saline was administered orally
methanol oxidation in rat hepatocytes. Free methanol is (75 mg/kg body weight), based on the earlier report (Leon
created from aspartame when it is heated to above 86°F et al. 1989). However, early reports on aspartame of higher
(30°C). This would occur when an aspartame-containing prod- doses included two different authors, namely, Labra-Ruiz et
uct is improperly stored or when it is heated (e.g. as part of a al. (2007) and Leon et al. (1989), and the reports are con-
‘food’ product such as Jello). Methanol is gradually released in troversial. This provided additional interest to use this dos-
the small intestine when the methyl group of aspartame age in our study. In order to confine within the human
encounters the enzyme chymotrypsin. Methanol breaks down exposure limit, this dose was selected. A 1 L (approx. 1
into formic acid and formaldehyde in the body. Formaldehyde quart) aspartame-sweetened beverage contains about 56 mg
is a deadly neurotoxin (Lee et al. 1994; Eells et al. 2000). of methanol was used. Heavy users of aspartame-containing
Rodents do not develop metabolic acidosis during methanol products consume as much as 250 mg of methanol daily, or
poisoning, owing to their high liver folate content, and in 32 times the EPA limit.
order to create similar results in human beings, only folate- MTX in sterile saline was administered (0.2 mg/kg/day)
deficient rodents are required to accumulate formate in order subcutaneously for 7 days to folate-deficient treated as well
to develop acidosis in methanol poisoning (Lee et al. 1994; as to folate-treated aspartame groups (Gonzalez-Quevedo
Eells et al. 2000). et al. 2002). One week after treatment with MTX, folate
Hence, in this study, in order to mimic the human deficiency was confirmed by estimating the urinary excretion
situation, a folate deficiency status was induced by ad- of formaminoglutamic acid (FIGLU) (Tabor and Wyngarden
ministering methotrexate (MTX). The focus of this study 1962). From the eighth day, only the MTX-treated aspartame
J. Biosci. 37(4), September 2012
Effect of aspartame on oxidative stress 681
group received the aspartame, whereas the other two groups for methanol was found to be 5 mg/100 mL and reproduc-
received equivalent volumes of saline as an oral dose and all ibility was>93%.
animals were handled similarly. The chronic dose of aspar-
tame was given for 90 days and all the animals were fed 2.7 Plasma corticosterone
folate-deficient diet except the control animals till 90 days.
Plasma corticosterone level was estimated according to the
2.4 Sample collections method described earlier by (Mattingly 1962) using excita-
tion at 470 nm and emission at 530 nm in fluorescence
The blood samples and isolation of brain was performed spectrophotometer.
between 8 and 10 a.m. to avoid circadian rhythm induced
changes. Stress-free blood samples were collected as per the 2.8 Lipid peroxidation
technique described by Feldman and Conforti (1980).
15 gm trichloro acetic acid was added to 100 mL of 0.25 N
hydrochloric acid. 15 mg of thio barbituric acid was dissolved
2.5 Brain dissection
in 4 mL of trichloro acetic acid in HCl mixture. To 0.1 mL of
homogenate, 0.4 mL trichloro acetic acid–thio barbituric acid–
The animals were sacrificed and the brain was immediately hydrochloric acid mixture was added and kept in a boiling
removed and washed with ice-cold phosphate buffered saline water bath for 20 min. Then, it was cooled to room temperature
(PBS). To expose the brain, the tip of curved scissors was gradually, followed by addition of 1 mL of n-butanol, vor-
inserted into the foramen magnum and a single lateral cut texed well and centrifuged for 10 min. The supernatant was
was made into the skull extending forward on the left and taken and was read at 532 nm in spectrophotometer. The
right side. With a bone cutter, the dorsal portion of cranium level of LPO was expressed as nanomoles of malondialde-
was peeled off, and using a blunt forceps, the brain was hyde (an intermediary product of lipid peroxidation, using
dropped onto the ice-cold glass plate, leaving the olfactory thio barbituric acid)/mg protein (Ohkawa et al. 1970).
bulbs behind. The whole process of removing brain took less
than 2 min. After removing the brain, it was blotted and
chilled. Further dissection was made on ice-cold glass plate. 2.9 Superoxide dismutase (EC 1.15.1.1) (SOD)
The discrete regions of brain (cerebral cortex, cerebellum,
midbrain, pons medulla, hippocampus and hypothalamus) 0.5 mL of homogenate was mixed with 0.5 mL of ethanol
were dissected according the method given by Glowinski chloroform mixture; the content was shaken well for 15 min
and Iverson (1996). The homogenate (10%w/v) of the indi- and the centrifuged for 10 min. To 0.5 mL of supernatant
vidual regions were prepared in a Teflon-glass tissue ho- taken, 2 mL of Tris-HCl buffer (pH 8.2) was added. Finally,
mogenizer, using ice-cold Tris HCl (100 mm, pH 7.4) buffer 0.5 mL of freshly prepared pyrogallol solution was added
and centrifuged separately in refrigerated centrifuge at 300g and read at 470 nm in 0, 1, 2 and 3 min intervals. The
for 15 min. The supernatant was used for analysing the activity was expressed as Units/min/mg protein (Marklund
parameters in this study. and Marklund 1974).
2.10 Catalase (EC.1.11.1.6)(CAT)
2.6 Blood methanol measurement
Four sets of 0.1 mL of homogenate was mixed with 1 mL of
100 mL plasma was deproteinized with equal volume of
phosphate buffer and 0.5 mL of H2O2, and this reaction was
acetonitrile and centrifuged for 7 min at 4°C (Dorman et al.
arrested by addition of 2 mL potassium dichromate acetic
1994). The supernatant (20 mL) was analysed for blood
acid reagent at various intervals of 0, 15, 30 and 60 s. Then
methanol and formate using a HPLC refractive index detec-
all the samples were kept in a boiling water bath for 10 min
tor system (Shimadzu RID, Japan) (equipped with Rezex
and were cooled gradually. The developed green colour was
ROA-organic acid column 300 mm × 7.5 mm I.D.,
read at 610 nm. The activity was expressed amount of H2O2
Phenomenex) with the security guard cartridge (AJO 4490
utilized/min/mg protein (Sinha 1972).
Phenomenex). Column oven was used to maintain the tem-
perature at 60°C. The mobile phase was 0.026 N sulphuric
acid (Pecina et al. 1984). By using methanol as an external 2.11 Glutathione peroxidase (EC.1.11.1.9) (GPx)
standard, the recovery of methanol (HPLC grade) from
blood was found to be 92–96%. Linearity for methanol To three sets of test tubes 0.4 mL of phosphate buffer,
was found to be 5–500 mg/100 mL. The detector sensitivity 0.1 mL of sodium azide, 0.2 mL of standard GSH solution
J. Biosci. 37(4), September 2012
682 Ashok Iyyaswamy and Sheeladevi Rathinasamy
and 1 mL of H2O were sequentially mixed. To this 0.1 mL of when there was a significant ‘F’ test ratio. The level of signif-
H2O2 was added and the reaction in individual set was icance was fixed at p≤0.05.
arrested with 1 mL of 10% TCA at 0, 1.5, 3 min intervals
and then centrifuged for 10 min. To the 0.2 mL of superna-
3. Results
tant 1.8 mL of H2O followed by 1 mL of 5% TCA were
added and allowed to stand at room temperature for 20 min.
Then 4 mL of phosphate solution was added, followed by The data from various groups for the individual parameters
0.5 mL of DTNB. The optical density was taken at 412 nm are presented as bar diagram with mean ± STD.
within 5 min. The GPx activity is expressed as units/min/mg
protein (Rotruck et al. 1973). 3.1 Blood methanol level and plasma corticosterone level
The data for the blood methanol level and plasma cortico-
2.12 Reduced glutathione
sterone level after chronic exposure to aspartame adminis-
tration are given in figure 1. Even after chronic exposure, the
To 0.2 mL of homogenate, 1.8 mL of H2O, 1 mL of 5%
aspartame-treated MTX animals showed marked increase in
TCA were sequentially added and kept at room temperature
the blood methanol level (df 2, F=2789) and plasma corti-
for 20 min. Then, 4 mL of 0.3 M phosphate solution fol- costerone level (df 2, F=874.6) from controls as well as from
lowed by 0.5 mL of DTNB was added and mixed well. The the MTX-treated group.
optical density was taken at 412 nm within 5 min. from the
addition of DTNB. Reduced glutathione (GSH) was mea-
sured by its reaction with 5.5- dithiobis 2 nitro benzoic acid 3.2 Lipid peroxidation, superoxide dismutase
(DTNB), to form a compound that absorbs at 412 nm and catalase levels
(Moron et al. 1979). The level of GSH is expressed as μg
of GSH/mg of protein. The results are given in figure 2. The LPO levels in the
lipid peroxidation (LPO) MTX-treated animals did not differ
2.13 Protein thiol from those in the controls. However, the aspartame-treated
MTX animals showed a marked increase in the LPO level in
the entire brain regions studied, which included cerebral
To 1 mL of homogenate was added 1.5 mL of Tris-
cortex (df 2, F=182), cerebellum (df 2, F=125.8), midbrain
hydrochloric acid buffer (pH 8.2) which contained 0.02
(df 2, F=272.2), pons-medulla (df 2,F=206.5), hippocampus
ethylene di-amine tetra acetic acid . This was followed by
(df 2, F=187.4) and hypothalamus (df 2, F=235.7) from the
the addition of 0.1 mL of DTNB solution and 7.5 mL meth-
control as well as from the MTX-treated animals.
anol. The contents were mixed well in a vortex mixture and
The results are given in figure 3. The superoxide dismu-
than centrifuged at 3000g for 10 min. The colour developed
tase (SOD) activity in the MTX-treated animals did not
was read at 412 nm. The level of thiol was expressed as
differ from those in the controls. However, the aspartame-
μg/mg protein. Tissues were analysed for protein and
treated MTX animals showed a marked increase in the SOD
sulfhydryl concentration (Sedlack and Lindsay 1968).
activity as compared to control and MTX-treated animals, in
the cerebral cortex (df 2, F=50.5), cerebellum (df 2, F=
2.14 Statistical analysis 19.2), midbrain (df 2, F=46.5), pons-medulla (df 2, F=
90.3) hippocampus (df 2, F=106) and hypothalamus(df 2,
Statistical analysis was carried out using the SPSS statistical F=84.8) from. The results are given in figure 4. The CAT
package version 17.0. The results are expressed as mean ± STD activity in the MTX-treated animals did not differ from the
and the data were analysed by the one-way analysis of variance controls. However, the aspartame-treated MTX animals
(ANOVA) followed by Turkey’s multiple comparison tests showed a marked increase in the CAT activity as compared
Blood Methanol Level Plasma Corticosterone Level
ug/dl of plasma
Methanol level
25 100
*# *#
20 80
(mM)
15 60 *
10 40
5 * 20
0 0
control MTX Asp+MTX control MTX Asp+MTX
Figure 1. Methanol level (mM) in rat blood upon aspartame administration (75 mg/kg) using HPLC and plasma corticosterone level.
J. Biosci. 37(4), September 2012
Effect of aspartame on oxidative stress 683
nm of malondialdehyde / mg
LPO activity
8 * * *
* *
* # # #
6 # #
#
protein
4 * *
2
0
CC CB MB PM HP HY
Figure 2. Effect of aspartame (75 mg/kg body weight) on LPO activity (mmol/mg tissue) in rat brain discrete regions.
to control and MTX treated animals, in the cerebral cortex (df 2, F=35.4) and midbrain (df 2, F=89.9). In the cerebellum
(df 2, F=130.2), pons-medulla (df 2, F=51), hippocampus (df 2, F=83.6), pons-medulla (df 2, F=67.7), hippocampus (df
(df 2, F=169) and hypothalamus (df 2, F=289.8). In the 2, F=81.7) and hypothalamus (df 2, F =136.9), a marked
midbrain (df 2, F=196.7) and cerebellum (df 2, F =126), increase in GPx activity as compared control was seen, and
only a marked increase in CAT activity as compared to this remained similar to MTX-treated animals. The results are
control was seen and this remained similar to MTX-treated given in figure 6.The GSH levels in the MTX-treated animals
animals. did not differ from the controls. However, the aspartame-
treated MTX animals showed a marked decrease in the GSH
levels as compared to control and MTX-treated animals, in
3.3 Glutathione peroxidase, reduced glutathione the cerebral cortex (df 2, F =21.4), midbrain (df 2, F=28.7),
and protein thiol levels pons-medulla (df 2, F=23.1) and hippocampus (df 2, F=
14.62). However, in the hypothalamus (df 2, F=19.4) the
The results are given in figure 5. The GPx activity in the GSH level showed marked decrease as compared to controls
MTX-treated animals did not differ from the controls. but not the MTX-treated group. Moreover, in the cerebellum,
However, the aspartame-treated MTX animals showed a the GSH level remained similar to that in control and MTX-
marked increase in the GPx activity as compared to treated animals. The results are given in figure 7. The protein
control and MTX-treated animals, in tge cerebral cortex thiol levels in the MTX-treated animals did not differ from
SOD Activity
Units/min/mg protein
5 *
#
4 *
3 #
* *
* *
2 # #
# #
1
0
CC CB MB PM HP HY
Figure 3. Effect of aspartame (75 mg/kg body weight) on SOD activity (units/mg tissue) in rat brain discrete regions.
J. Biosci. 37(4), September 2012684 Ashok Iyyaswamy and Sheeladevi Rathinasamy
Amount of H2O2 utilized/
CAT activity
15 * *
min/ mg protein
* * * * #
#
# # # #
10
* *
5
0
CC CB MB PM HP HY
Figure 4. Effect of aspartame (75 mg/kg body weight) catalase activity (units/mg tissue) in rat brain discrete regions.
those in the controls. However, the aspartame-treated MTX blood methanol in the aspartame-treated animals. Methanol
animals showed a marked decrease in the protein thiol levels is metabolized by three enzyme systems, namely, the alcohol
as compared to control and MTX-treated animals, in the dehydrogenase system, the catalase peroxidative pathway
entire brain regions studied such as the cerebral cortex (df and the microsomal oxidizing systems. Among these, the
2, F=114.7), cerebellum (df 2, F=158.9), midbrain (df 2, F= microsomal oxidizing system is reported to be responsible
83.7), pons-medulla (df 2, F=37.1), hippocampus (df 2, F= for free radical generation (Goodman and Tephly 1968).
68.6) and hypothalamus (df 2, F=119.3). Exposure to methanol causes oxidative stress by altering
the oxidant/antioxidant balance in lymphoid organs of rat
(Parthasarathy et al. 2006). The condition could also be
4. Discussion associated with the abundance of redox active transition
metal ions, and the relative death of antioxidant defence
This study deals with strengthening the toxic metabolite system (Samuel et al. 2005). In addition to that, oxidative
methanol released in the body after the consumption of stress may lead to the generation of superoxide, peroxyl and
aspartame. The alteration in the free-radical-scavenging hydroxyl radicals (Keyhani and Keyhani 1980).The increase
enzymes in the aspartame-administered animals clearly indi- in free radicals could not be ignored as cells can be injured or
cates that free radical generation may be due to the methanol killed when the ROS generation overwhelms the cellular
which is one of the metabolic products of aspartame. Even antioxidant capacity (Oberly and Oberly 1986).
after chronic aspartame administration, there was detectable Particularly, the brain is more vulnerable to oxidative
GPx Activity
*
* * *
units/min/mg protein
* # *
10 # # # # #
8 * * *
6
4
2
0
CC CB MB PM HP HY
Figure 5. Effect of aspartame (75 mg/kg body weight) on GPx activity (units/mg tissue) in rat brain discrete regions.
J. Biosci. 37(4), September 2012Effect of aspartame on oxidative stress 685
Reduced GSH Level
µg of GSH/mg of
0.06 * *
0.05 # *
*
# *
protein
0.04 #
0.03 #
*
0.02
0.01
0
CC CB MB PM HP HY
Figure 6. Effect of aspartame (75 mg/kg body weight) on GSH concentration (mmol/mg tissue) in rat brain discrete regions.
damage due its high oxygen consumption and due to the LPO is a free-radical-mediated process. In the entire rat
presence of high levels of polyunsaturated fatty acids (Floyd brain regions after aspartame consumption, a marked in-
and Carney 1992). crease in LPO was noted, which also supports the generation
SOD converts superoxide anion (O2−) to hydrogen per- of free radicals. Generally, when the generation of reactive
oxide (H2O2) (Akyol et al. 2002), which are subsequently free radicals overwhelms the antioxidant defense, LPO of the
converted to water and molecular oxygen by glutathione cell membrane occurs. In spite of the increase in the SOD,
peroxidase or catalase (Dringen 2000). GPx is a major anti- CAT and GPx enzyme system in order to prevent the accu-
oxidant enzyme in many tissues and has been speculated to mulation of free radical, a marked increase in the LPO in the
be a major antioxidative mechanism in the brain (Barlow- entire brain regions indicated this result and possible loss of
Walden et al. 1995). In this study there was a significant membrane integrity. Moreover, reactive-oxygen-mediated
increase in the SOD activity after chronic aspartame inges- damage could not be overlooked particularly to proteins,
tion with an associated increase in the catalase activity. As which has been shown to deleteriously affect cellular func-
catalase is the main scavenger of H2O2 at high concentration tions (Sohal et al. 2002).
(Kono and Fridorich 1982), that indicates the accumulation GSH is considered to be one of the most important com-
of H2O2 as a result of increased SOD activity. It has been ponents of the antioxidant defense of living cells. The re-
found that consumption of methanol provokes changes in the duced tri-peptide GSH is a hydroxyl radical and singlet
activity of antioxidant enzymes with an increase in the oxygen scavenger, and participates in a wide range of cellu-
activity of catalase (Skrzydlewska et al. 1998). Hence, the lar functions. Since GSH is present in high intracellular
increase in this enzyme activity may also be due to the free concentrations, there is a high probability that reactive
radicals generated. According to Yu (1994), free radicals oxygen species (such as superoxide, singlet oxygen and
may also induce the expression of antioxidant enzymes, hydrogen peroxide) is to be quenched by reaction with
thereby enhancing the neuronal resistance to subsequent GSH before they can initiate their chain reaction-
oxidative challenges. damaging effects (Jones et al. 1981). The depletion of
Protein Thiol Level
mmoles / mg protein
8 control
6 * * ** ** **
* * * MTX
4 # # # # *
#
#
2 Asp+MTX
0
CC CB MB PM HP HY
Figure 7. Effect of aspartame (75 mg/kg body weight) on protein thiol levels (mmol/mg tissue) in rat brain discrete regions. CC, cerebral cortex;
CB, cerebellum; MB, mid brain; PM, pons medulla; HI, hippocampus; HY, hypothalamus. The level of significance was fixed at p≤0.05 (n= 6).
J. Biosci. 37(4), September 2012686 Ashok Iyyaswamy and Sheeladevi Rathinasamy
GSH may be one of the reasons for the increase in the Acknowledgements
vulnerability added to free-radical-induced damage. In addi-
tion, the decrease in GSH concentration is quite possible The author is grateful for the valuable suggestion offered by
because the methanol metabolism also depends upon GSH Dr NJ Parthasarathy and Dapkupar Wankhar. The financial
(Pankow and Jagielki 1993). assistance provided by the University of Madras is gratefully
The significant decrease in the protein thiols observed in acknowledged.
this study may be due to the oxidation of proteins in oxida-
tive process, and it is also justified by the decreased level of
one of the major thiol substance GSH. Similarly decreased References
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MS received 18 April 2012; accepted 06 June 2012
Corresponding editor: INDRANEEL MITTRA
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