The Scandinavian Journal of Clinical & Laboratory Investigation,

Vol. 69, No. 3, May 2009, 371379

ORIGINAL ARTICLE
Amelioration of streptozotocin-induced diabetes mellitus, oxidative stress and
dyslipidemia in rats by tomato extract lycopene
Mamdouh M. Ali1 and Fatma G. Agha2
1
Department of Biochemistry, Division of Genetic Engineering and Biotechnology, National Research Centre, Cairo, Egypt;
2
Department of Forensic Medicine and Toxicology, Faculty of Medicine for Girls, El-Azhar University, Cairo, Egypt

The effects of various doses of lycopene were studied in streptozotocin (STZ)-induced hyperglycaemic rats to
evaluate its possible hypoglycaemic, hypolipidaemic and antioxidant activity in diabetes. Compared to the
Scand J Clin Lab Invest Downloaded from informahealthcare.com by Boston University on 07/24/13

normoglycaemic group, the treatment of rats with a single dose of STZ (65 mg/kg body weight) revealed a
significant increase (pv0.05) only in plasma hydrogen peroxide (H2O2), i.e. by 230 %; it increased the
thiobarbituric acid reactive substances (TBARS) as index of the lipid peroxidation level by 69 %, while total
antioxidant activity was decreased by 36 %, with a consistently significant decrease (pv0.05) in the activity of
erythrocytes antioxidative enzymes catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase
(GPx). The levels of total lipid, triglycerides and total cholesterol in serum of hyperglycaemic rats were increased
by 14 %, 65 % and 36 %, respectively, while HDL-C decreased by 22 % compared to the normoglycaemic group.
Exogenous administration of individual gradual doses of lycopene to hyperglycaemic rats causes a dose-
dependent decrease in glucose level, an increase of insulin concentration, a decrease of H2O2 and TBARS levels,
as well as increased total antioxidant status with increased antioxidant enzyme activities (CAT, SOD and GPx)
with improvement in serum lipid profile. It is obvious from this study that lycopene acts as an antidiabetic agent
through lowering the free radical and has an improving effect on serum that reaches the normal level; the greatest
effect of lycopene was observed at 90 mg/kg body weight.
For personal use only.

Keywords: Antioxidative enzymes; hyperglycaemia; lipid peroxidation; lipid profile; lycopene

Introduction agents continues to be an important area for research.

Diabetes mellitus (DB) is one of the most crippling
diseases that man has ever had to deal with, and its
prevalence has risen dramatically over the past two
decades [1]. Currently, there are over 150 million
diabetics worldwide, and this is likely to increase to
300 million or more by the year 2025 due to increased
sedentary lifestyle, consumption of energy-rich diet
and obesity [2]. Type II DM (T2DM) develops when
the pancreas cannot produce enough insulin or when
the body tissue becomes resistant to insulin, and
without insulin the body cannot process glucose,
resulting in too much sugar in the blood and not
enough in the bodys cells [1]. This disturbance
increases the risk of many disorders, including obesity,
hypertension, hyperuricaemia and dyslipidaemia [3,4].
Synthetic antidiabetic agents used in the treatment
of diabetes can produce serious side effects, including
hypoglycaemic coma and disturbances of the liver and
kidneys. In addition, the majority of these agents are
not suitable for use in pregnant women [5]. Therefore,
the search for more effective and safer antidiabetic
Following the recommendations of the World Health
Organization [6] on the beneficial uses of medicinal
plants in the treatment of DM, investigation of
antidiabetic agents from medicinal plants has also
become more important.
It has been reported that diabetic patients have
significant defects of antioxidant protections and
generation of reactive oxygen species (oxidative
stress) which may be important in the aetiology of
diabetic complications [7]. Decreased superoxide
dismutase (SOD), catalase (CAT), ceruloplasmin
and glutathione peroxidase (GPx) activities, as well
as a decrease in the GSH level and an increase in the
concentration of glutathione disulphide (GSSG),
were observed in erythrocytes of diabetic patients
and in tissues from diabetic animals [8].
Lipid peroxidation products, which increase in
clinical and experimental diabetes, are important
results of oxygen-derived free radicals stress. These
products may be important in the pathogenesis of
vascular complications in DM [9]. Nitric oxide (NO)

Correspondence: Mamdouh M. Ali, National Research Center, Biochemistry Department, Division of Genetic Engineering and Biotechnology, El Tahrir St., El
Dokki 12622, Cairo, Egypt. Fax: +00202 33370931. Email: mmali1999@yahoo.com

(Received 13 June 2008; accepted 12 November 2008)

ISSN 0036-5513 print/ISSN 1502-7686 online # 2009 Informa UK Ltd (Informa Healthcare, Taylor & Francis AS).
DOI: 10.1080/00365510802658473
 372 M. M. Ali & F. G. Agha
is considered a potent endothelium-derived vasodi-
lator that participates in the general homeostasis of
the vasculature. Previous studies have demonstrated
that the development of diabetic complications in
diabetes is closely related to the increased generation
of superoxide anion (O2N) and NO [10].
Several natural agents have been used to amelio-
rate some toxic and carcinogenic xenobiotics and
drug toxicity. These include vitamins E and C, garlic
extract and carotenoids [1114]. Carotenoids are a
family of fat-soluble pigments found in tomatoes and
its products, some other fruits and vegetables, and
many studies have investigated their potential in
oxidative stress. Tomato carotenoids include lyco-
Scand J Clin Lab Invest Downloaded from informahealthcare.com by Boston University on 07/24/13
pene and other similar carotenoids [15,16].
Lycopene, a carotenoid compound occurring natu-
rally in tomato and other fruits, has long been known for
its health-promoting role in the prevention of chronic
diseases such as cancer and cardiovascular diseases [17].
However, the biochemical mechanisms underlying the
health-promoting roles are not fully understood,
although the antioxidative activity of lycopene [18],
which has been shown to be a potent protector against
oxidative damage to DNA, protein and lipids, is thought
For personal use only.
to be primarily responsible. Other activities of lycopene,
such as modulation of cellcell communication [19],
inhibition of cell proliferation [20], resistance to bacterial
infections [21], may also be involved.
Although the previous beneficial effects of lycopene
have been established in various disorders, little atten-
tion has been given to the role of lycopene as antidiabetic
and hypolipidaemic agents. The present study was
undertaken to investigate the effect of lycopene on
streptozotocin (STZ)-induced hyperglycaemic rats to
evaluate the possible hypoglycaemic, antioxidant and
hypolipidaemic activity of lycopene in diabetes as well as
serum amylase activity in diabetes mellitus.
Material and methods
Animals
Animal care and handling was in accordance with the
guidelines set by the World Health Organization,
Geneva, Switzerland. With the approval of the
committee for animal care at the National Research
Centre, Egypt, healthy adult male Sprague-Dawley
rats with an average body weight of 17020 g were
obtained from the animal house of the National
Research Centre and used in the study. The animals
were housed under standard laboratory conditions
(12 h light and 12 h dark) in a room with controlled
temperature (243 C) during the experimental per-
iod. The rats were provided ad libitum with tap water
and fed with standard commercial rat chow.
Experimental design
Rats were divided into two experimental groups. (A)
The normoglycaemic group received one dose (1 mL)
via intraperitoneal (i.p.) injection of 0.1 M citrate
buffer, pH 4.5. (B) The hyperglycaemic group was
induced to develop diabetes mellitus by a single i.p.
injection of the diabetogenic agent streptozotocin
(STZ; Sigma Chemical Co., St. Louis, Mo., USA) in
a dose of 65 mg/kg body weight. STZ was dissolved
in 1 mL of 0.1 M citrate buffer pH 4.5 [22]. After
72 h, tail blood samples were obtained from each rat,
and the blood glucose concentration was determined
spectrophotometrically using Roche Diagnostics kits
(Germany). STZ-treated rats with blood glucose
below 300 mg/dL were excluded from the study [23].
Each group (A and B) was divided into two
subgroups. The control subgroup received 0.5 mL
corn oil by oral gavage. Lycopene (Sigma Chemical
Co., St. Louis, Mo., USA) was suspended in corn oil
and administered to animals by oral gavage at doses of
10, 30, 60 or 90 mg lycopene/kg body weight for 30
consecutive days. The dose of lycopene used in this
study was selected on the basis of previous studies [24].
Twenty-four hours after the previous dose, the
rats were anaesthetized by i.p. injection with sodium-
5-ethyl-5-(1-methyl-butyl)-2-thiobarbiturate
(Thiopental) (VUAB, Roztoky, Czech Republic) at a
dose of 35 mg/kg body weight and then killed by
cervical dislocation. Blood samples were collected
from the anaesthetized rats, by puncture of the right
ventricle, in heparinized tubes and centrifuged for
15 min at 1000g. Plasma was carefully removed. The
separated cells were then washed three times and
resuspended in a 0.9 % NaCl solution and repeated
centrifugation. The washed cells were lysed in an
equal volume of water and mixed thoroughly. The
haemoglobin concentration was estimated in an
aliquot of the hemolysate using a commercial assay
kit (Sigma Chemical Co., St. Louis, Mo., USA). The
hemolysate was used for estimation of catalase
(CAT), superoxide dismutase (SOD) and glutathione
peroxidase (GPx) activities. While plasma used in the
assay the level of glucose, insulin, thiobarbituric acid
reactive substance (TBARS), hydrogen peroxide
(H2O2), total antioxidant status and nitric oxide
(NO). In addition, pancreas were collected and stored
at 280 C until the time of analysis.
Biochemical analysis
Determination of insulin concentration
The concentration of insulin in plasma was estimated
using an ELISA kit (Mercodia Ultrasensitive Mouse
Insulin, Sweden), which is also suitable for determining

insulin in rats. It is based on the direct sandwich
technique in which two monoclonal antibodies are
directed against separate antigenic determinants on the
insulin molecule. The reaction is read spectrophoto-
metrically at 450 nm. The coefficient of variation is
12 %, with lowest detectable border of 0.2 ng/mL.
Total pancreatic insulin content
The pancreas was weighed and then homogenized in
acid ethanol (75 % ethanol, 23.5 % distilled water
and 1.5 % concentrated HCl) at 10 mL/g of tissue.
After overnight incubation at 4 C, the suspensions
were centrifuged at 2000g for 10 min and the super-
Scand J Clin Lab Invest Downloaded from informahealthcare.com by Boston University on 07/24/13
natants collected and analysed for insulin content
using an ELISA kit (Mercodia Ultrasensitive Mouse
Insulin, Sweden) [25]. The coefficient of variation was
12 %, with lowest detectable border of 0.2 ng/mg
tissue.
Determination of hydrogen peroxide (H2O2) level
The level of plasma H2O2 was determined by the
method of Wolf [26] based on ferrus oxidation with
xylenol orange (Fox-1) reagent and colour develop-
For personal use only.
ment being read spectrophotometrically at 560 nm.
The coefficient of variation was 14 %, with lowest
detectable border of 1.0 mmol/L.
Assessment of lipid peroxidation
As a marker of lipid peroxidation, thiobarbituric acid
reactive substances (TBARS) were measured using
the method of Conrad et al. [27], based on the
reaction of thiobarbituric acid with malondialdehyde,
the second products of lipid, carbohydrate, protein
and DNA peroxidation, as follows: a 0.5 mL sample
was shaken with 2.5 mL of 20 % TCA. One millilitre
of 0.67 % thiobarbituric acid was added to the
mixture, which was shaken and kept for 30 min in a
boiling water bath followed by rapid cooling. Then
4 mL of n-butyl alcohol was added and shaken. The
mixture was centrifuged at 3000 rpm for 10 min. The
resultant n-butyl alcohol layer was taken into a
separate tube and the TBARS content was deter-
mined calorimetrically from the absorbance at
535 nm. 1,1,3,3,-tetra-ethoxypropane was used as
standard. Level of peroxidation products was
expressed as the amount of TBARS in plasma
(nmol/mL). The coefficient of variation is 16 %, with
lowest detectable border of 1.0 nmol/mL.
Determination of total antioxidant status
Antioxidant status was assayed with a Randox kit
purchased from Randox Laboratories Ltd (Antrim,
Effect of lycopene on hyperglycaemia 373
UK). The assay is based on the reaction of
metmyoglobin with hydrogen peroxide to form
ferrmyoglobin, a free radical species. A chromogen
2,2-amino-di-(3-ethylbenzenthiazole sulphate) is
incubated with ferrmyoglobin to produce a radical
cation which has a relatively stable blue colour.
Antioxidants in the added plasma can suppress this
colour production to a certain degree proportionally
to their concentrations; the intensity of blue colour
was measured at 600 nm. The coefficient of variation
is 11 %, with lowest detectable border of 1.1 nmol/
mL.
Determination the level of nitric oxide (NO)
NO was determined according to the methods of
Granger et al. [28] based on NO production being
detected in biological fluids via nitrite using a Roche
nitrite/nitrate assay kit. The nitrate present in the
sample is reduced to nitrite by reduced nicotinamide
adenine dinucleotide phosphate (NADPH) in the
presence of the enzyme nitrate reductase. The nitrite
formed reacts with sulphanilamide and N-(1-
naphthyl)-ethylenediamine dihydrochloride to give a
complex which measured 550 nm. The coefficient of
variation is 15 %, with lowest detectable border of
3.0 mmol//mL.
Determination of catalase (CAT) activity
CAT activity was adapted from the method of Aebi
[29], based on the principle that the disappearance of
hydrogen peroxide was monitored spectrophotome-
trically at 240 nm. Briefly, 10 mL of the hemolysate
was added to a cuvette containing 2.89 mL of
50 mmol/L potassium phosphate buffer (pH 7.4),
the reaction initiated by adding 0.1 mL of 30 mmol/
L H2O2 to make a final volume of 3.0 mL at 25 C.
The decomposition rate of H2O2 was measured at
240 nm for 5 min using a spectrophotometer.
Activity was defined as the mmol H2O2 decreased/
min/g Hb. The coefficient of variation is 13 %, with
lowest detectable border of 7.0 mmol /min/g Hb.
Determination of superoxide dismutase (SOD) activity
SOD activity was measured spectrophotometrically
following a method developed by Marklund &
Marklund [30] in which colour change by the auto-
oxidation of pyrogallol is used. Briefly, 100 mL of the
hemolysate was mixed with 1.5 mL of a Tris-EDTA-
HCl buffer (pH 8.5); 100 mL of 15 mmol/L pyrogal-
lol was added and the reaction mixture was incubated
at 25 C for 10 min. The reaction was terminated by
adding 50 mL of 1 N HCl and measurements were
 374 M. M. Ali & F. G. Agha
taken at 440 nm. One unit was determined as the
amount of enzyme that inhibited the oxidation of
pyrogallol by 50 %. The activity was expressed as
units/g Hb. The coefficient of variation is 12 % lowest
detectable border of 70.0 unit/g Hb.
Determination of glutathione peroxidase (GPx) activity
GPx activity was measured using the Paglia &
Valentine method [31], where the oxidation of
NADPH by hydrogen peroxide was followed at
340 nm at 25 C. Briefly, the reaction mixture con-
tained 2.525 mL of 0.1 mol/L Tris-HCl buffer
(pH 7.2), 75 mL of 30 mmol/L glutathione, 100 mL
Scand J Clin Lab Invest Downloaded from informahealthcare.com by Boston University on 07/24/13
of 6 mmol/l NADPH and 100 mL of glutathione
reductase (0.24 unit). One hundred microlitres of the
hemolysate was added to 2.8 mL of the reaction
mixture and incubated at 25 C for 5 min. The reaction
was initiated by adding 100 mL of 30 mmol/L H2O2
and the absorbance was measured at 340 nm for
5 min. Activity was expressed as the oxidized NADPH
nmol/g Hb. The coefficient of variation was 12 %, with
lowest detectable border of 3.0 nmol/g Hb.
For personal use only.
Assessment of lipid profile
Lipid components, including serum total lipid,
triglycerides, total cholesterol, HDL-cholesterol
(HDL-C) and LDL-cholesterol (LDL-C), were mon-
itored by enzymatic methods using commercial kits
obtained from BioMeriux (France). The coefficients
of variation are 14 %, 13 %, 22 %, 17 % and 13 %,
respectively, with lowest detectable borders of 230.0,
26.0, 51.0, 21.0 and 27.0 mg/dL, respectively.
Statistical analysis
Data were expressed as meanstandard error (S.E.)
and analysed statistically using a one-way analysis of
Table I. The effects of various doses of lycopene on levels of plasma glucose and insulin, as well as the total pancreatic insulin
content of the normoglycaemic and hyperglycaemic rats.
Lycopene (mg/kg body wt)
Control 10
Glucose (mg/dL)
Normoglycaemic 80.57.5 82.77.4
Hyperglycaemic 370.025.4a 375.013.6a
Insulin (ng/mL)
Normoglycaemic 5.20.7 5.60.8
Hyperglycaemic 1.90.2a 1.80.3a
Pancreatic insulin (ng/mg tissue)
Normoglycaemic 32.03.0 35.03.8
Hyperglycaemic 19.00.2a 19.71.4a
a
Significant difference from the normoglycaemic group. bSignificant difference from the hyperglycaemic group.
variance (ANOVA) followed by Students t-test for
comparison between groups. Differences were con-
sidered significant at pv0.05.
Results
Effect of lycopene on plasma glucose and insulin levels
as well as the pancreatic insulin content
The plasma glucose level (Table I) in the hypergly-
caemic group was increased by 360 % (from
80.57.5 to 370.025.4 mg/dL; pv0.05) in compar-
ison to the normoglycaemic group. The administra-
tion of gradual doses of lycopene of 10, 30, 60 or
90 mg/kg body weight to rats with hyperglycaemia
resulted in decreased glucose concentrations by 0, 46,
65 and 78 %, respectively.
The plasma insulin level (Table I) in the hyper-
glycaemic group was decreased by 63.5 % (from
5.20.6 to 1.90.2 ng/mL; pv0.05) compared to
the normoglycaemic group. The administration of
gradual doses of lycopene to rats with hyperglycae-
mia resulted in increased insulin concentration (by 0,
105, 110 and 158 %, respectively).
The pancreatic insulin content (Table I) in the
hyperglycaemic group was decreased by 41 % (from
32.03.0 to 19.01.2 ng/mg tissue; pv0.05) com-
pared to content in the normoglycaemic group. The
administration of gradual doses of lycopene to rats
with hyperglycaemia resulted in increased insulin
concentration by 0, 20, 34 and 48 %, respectively.
Effect of lycopene on plasma H2O2, TBARS, total
antioxidant and NO levels
The level of H2O2 (Figure 1) in plasma of rats with
hyperglycaemia was increased by 326 % (from
1.90.3 to 8.30.6 mmol/L; pv0.05) compared to
the level in the normoglycaemic group. The TBARS
30 60 90
83.79.5 84.39.3 70.78.4
200.015.4a,b 130.017.5a,b 81.68.6a,b
4.40.5 5.50.7 5.70.7
3.90.5a,b 4.00.2a,b 4.90.5b
35.03.5 36.54.6 38.53.7
22.83.3a,b 25.53.2a,b 28.23.3a,b
 Effect of lycopene on hyperglycaemia 375

10 8
9 * *
Normoglycaemic 7 Normoglycaemic
Hyperglycaemic Hyperglycaemic
8
6

TBARS (nmol/ml)
7
H2O2 (mol/L)

*# 5 *#
6
*# *#
5 4 *#
4 *# 3
3
2
2
1 1
0 0
0 20 40 60 80 100 0 20 40 60 80 100
Scand J Clin Lab Invest Downloaded from informahealthcare.com by Boston University on 07/24/13

Dose of lycopene mg/Kg body weight Dose of lycopene mg/Kg body weight

5.5 34
5.0 32 Normoglycaemic
*# 30 * Hyperglycaemic
4.5
Total antioxidant (nmol/L)

28
4.0 26 *#
24
3.5 NO (mol/ml) 22
*# 20
3.0 18 *#
2.5 16
*# 14
2.0
For personal use only.

* 12 *#
1.5 10
8
1.0 Normoglycaemic 6
0.5 Hyperglycaemic 4
2
0.0 0
0 20 40 60 80 100 0 20 40 60 80 100
Dose of lycopene mg/Kg body weight Dose of lycopene mg/Kg body weight

Figure 1. Levels of plasma H2O2, TBARS, total antioxidant and NO in the normoglycaemic and hyperglycaemic rats after
administration of different concentrations of lycopene (10, 30, 60 or 90 mg/kg body weight). *Significant difference from the
normoglycaemic group. #Significant difference from the hyperglycaemic group.

level in rats treated with STZ was increased by 128 % (from 19.52.30 to 9.71.3 mmol/min/gHb;
(from 2.90.3 to 6.600.5 nmol/mL; pv0.05), while pv0.05), 54 % (from 380.233.6 to
total antioxidant was decreased by 75 % (from 172.919.4 unit/g Hb; pv0.05) and 35 % (from
2.80.4 to 1.60.2 nmol/L; pv0.05), and NO was 6.50.5 to 4.20.4 mmol/ min/gHb; pv0.05),
increased by 186 % (from 8.80.9 to 25.24.1 mmol/ respectively, compared to the results obtained from
mL; pv0.05) compared to the normoglycaemic group. the normoglycaemic group.
Compared to the hyperglycaemia untreated group, Compared to the hyperglycaemia untreated
administration of gradual doses of lycopene to rats group, administration of gradual doses of lycopene
with hyperglycaemia resulted in a decrease of the H2O2 to rats with hyperglycaemia resulted in increased
level by 14.5, 24, 40, 61 %, respectively, and the level of CAT activity by 8, 38, 85 and 150 %, respectively,
TBARS by 16, 38, 43, 50 %, respectively, while the increased SOD activity by 25, 58, 120 and 184 %,
level of total antioxidant activity was increased by 19, respectively, and increased GPx activity by 17, 25, 50
37.5, 81 and 160 %, respectively, and the level of NO and 86 %, respectively.
by 12, 19, 44 and 62 %, respectively.
Effect of lycopene on the lipid profile
Effect of lycopene on the activity of erythrocytes CAT, The levels of total lipid, triglycerides and total
SOD and GPx enzymes cholesterol (Table II) in serum of hyperglycaemic
The activities of CAT, SOD and GPx (Figure 2) in rats were increased by 14 %, 65 % and 36 %,
erythrocytes of rats receiving STZ decreased by 50 % respectively, while HDL-C decreased by 22 % com-
 376 M. M. Ali & F. G. Agha

34 total lipid levels by 5, 11, 16 and 19 %, respectively;

32
30 decreased levels of triglyceride by 5, 13, 33 and 57 %;
CAT (mol min/gHb)

28 *# decreased levels of total cholesterol by 16, 29, 32 and

26
24 37 %; increased levels of HDL-C by 20, 33, 50 and 82;
22
20 # and decreased levels of LDL-C by 1, 11, 17, and 25%.
18
16
14 #
12 * Discussion
10
8 DM and its sequels, neuropathy and angiopathy, are
6 Normoglycaemic
4 conditions in which oxidant-free radicals are involved
2 Hyperglycaemic
0
in both human and experimental models [32,33].
0 20 40 60 80 100 Several mechanisms may lead to increased oxidative
Dose of lycopene mg/Kg body weight stress in diabetes. First, hyperglycaemia may increase
the generation of free radicals through the ability of
Scand J Clin Lab Invest Downloaded from informahealthcare.com by Boston University on 07/24/13

600
glucose to enolize and yield oxidizing intermediates
550 *#
such as superoxide anion, hydroxyl radical, hydrogen
500
* * peroxide and nitric oxide [34,35]. Second, antioxidant
450
#
SOD (unit/gHb)

400
defences are reduced in diabetes [36]. Hence, com-
350 pounds with both antidiabetic and anti-oxidative
300 *# properties would be useful anti-diabetic agents [37].
250 ROS function as signalling molecules to activate
200 * several stress-sensitive pathways. In type II diabetes,
150 there is growing evidence that activation of stress-
100 sensitive pathways, such as nuclear factor-kB (NF-
Normoglycaemic
50 kB), NH2-terminal Jun kinases/stress activated pro-
For personal use only.

Hyperglycaemic
0 tein kinases (JNK/SAPK), p38 mitogen-activated
0 20 40 60 80 100
protein (MAP) kinase and hexosamine, are linked
Dose of lycopene mg/Kg body weight
to insulin resistance and b-cell dysfunction [38].
A relationship between diabetes and oxidative
10 stress has been confirmed in both experimental STZ
9 and human DM, together with reduced total anti-
*#
8 oxidant defence and depletion in individual antiox-
GPx (mol/min/gHb)

# idants [39]. Such a pro-oxidant environment was

7
demonstrated in our study by the increased level of
6 # lipid peroxidation products, nitric oxide and hydro-
5 * gen peroxide accompanied by inhibition of the
4 activity of antioxidative enzymes CAT, SOD and
3 GPx as well as total antioxidant status. Our results
2 were in agreement with those reported by Piconi et al.
Normoglycaemic [40], who mentioned that in diabetes overproduction
1
Hyperglycaemic of ROS can lead to a decrease in cell/organism
0
0 20 40 60 80 100
antioxidant defences, demonstrated as a fall in the
Dose of lycopene mg/Kg body weight concentration of a single antioxidant molecule or in
total antioxidant status. An increase in TBARS was
Figure 2. The activities of CAT, SOD and GPx in the presumably associated with an increase in ROS,
erythrocytes of normoglycaemic and hyperglycaemic rats confirming the fact that the singlet oxygen and
after administration of different doses of lycopene (10, 30, peroxyl radicals inhibited the activity of SOD [41].
60 or 90 mg/kg body wt). *Significant difference from the
Also Blum & Fridovich [42] mentioned that over-
normoglycaemic group. #Significant difference from the
hyperglycaemic group. production of superoxide radicals during oxidative
stress could inhibit the activity of GPx, while the
pared to the level in the normoglycaemic group; activity of CAT was inhibited by production of
however, the level of LDL-C did not change. singlet oxygen, superoxide and peroxyl radicals
Compared to the hyperglycaemia untreated [41,43]. In addition, Sowers [44] mentioned that high
group, the administration of gradual doses of glucose level induced inhibition of IL-1b-stimulated
lycopene to hyperglycaemic rats resulted in decreased NO release and NO synthase expression.
 Effect of lycopene on hyperglycaemia 377

Table II. The values of serum total lipid (TL), triglycerides (TG), cholesterol, HDL-cholesterol (HDL-C) and LDL-
cholesterol (LDL-C) in the normoglycaemic and hyperglycaemic rats after administration of lycopene.

Lycopene (mg/kg body wt)

Control 10 30 60 90

TL (mg/dL)
Normoglycaemic 250.330.3 268.231.9 250.733.4 242.420.7 235.327.4
Hyperglycaemic 285.731.3a 270.529.7a 253.431.7b 240.028.1b 230.426.3a,b
TG (mg/dL)
Normoglycaemic 38.63.3 37.72.6 35.93.2 40.23.4 25.54.9
Hyperglycaemic 63.69.2a 60.27.8a 55.37.0a,b 42.83.6b 27.14.3a,b
Cholesterol (mg/dL)
Normoglycaemic 65.07.3 64.28.5 65.78.6 63.06.6 51.27.6
88.28.9a 74.28.4a 62.26.3b 60.37.2b 55.26.3a,b
Scand J Clin Lab Invest Downloaded from informahealthcare.com by Boston University on 07/24/13

Hyperglycaemic
HDL-C (mg/dL)
Normoglycaemic 26.83.5 25.93.1 26.92.6 24.22.6 29.12.9
Hyperglycaemic 20.82.7a 24.93.4 27.63.5b 31.33.7b 37.84.6a,b
LDL-C (mg/dL)
Normoglycaemic 30.53.7 29.43.3 28.73.6 29.52.6 29.23.1
Hyperglycaemic 30.64.2 30.94.2 27.22.7b 25.32.7b 22.83.3a,b
a
Significant difference from normoglycaemic group. bSignificant difference from hyperglycaemic group.

In our study, lipid profile in the hyperglycaemic lycopene molecule, converting it to the energy-rich
For personal use only.

rats treated with STZ was disturbed compared to that
of the normoglycaemic group. Our result confirms
the previous data reported by Gary & Grundy [45],
and can be explained by the fact that type II DM is a
major risk factor for coronary artery disease. Patients
with diabetes have elevated LDL and VLDL
cholesterols and decreased HDL cholesterol [46,47].
In addition, Laakso [48] reported that type II
diabetes was associated with obesity, insulin resis-
tance, hypertension, high triglyceride and low HDL,
increasing the risk of cardiovascular disease.
Lamarche et al. [49] mentioned that a central
characteristic of dyslipidaemia in patients with type
II diabetes is an elevated triglyceride level, particu-
larly triglyceride-rich VLDL levels and decreased
HDL cholesterol levels. In diabetic patients, the
concentration of LDL cholesterol is usually not
significantly different from that seen in non-diabetic
individuals. However, patients with type II diabetes
typically have a preponderance of smaller, denser,
oxidized LDL particles, which may increase ather-
ogenicity even if the absolute concentration of LDL
cholesterol is not elevated.
Lycopene is a naturally present carotenoid in
tomatoes. It is an open-chain hydrocarbon contain-
ing 11 conjugated and 2 non-conjugated double
bonds arranged in a linear array [50]. Because of
the high number of conjugated dienes of lycopene, it
is the most potent singlet oxygen (1O2) quencher
among the natural carotenoids [51]. During 1O2
quenching, energy is transferred from 1O2 to the
triplet state. Trapping of other ROS, like OH, NO or
peroxynitrite, in contrast, leads to oxidative break-
down of the lycopene molecules. Thus, lycopene may
protect lipids, proteins and DNA against oxidation
[16,52].
The present study suggests that exogenous admin-
istration of different doses of lycopene to hypergly-
caemic rats causes a dose-dependent decreased level
of hydrogen peroxide, lipid peroxidation and NO as
well as increased activity of antioxidant enzyme.
These results indicate the important role of lycopene
in the reduction of oxidative stress. However, the
mechanism by which lycopene could affect free
radical and peroxide production, as well as total
antioxidant, CAT, SOD and GPx activities, is due to
its highly efficient antioxidant with a singlet-oxygen
and free radical scavenging capacity [53,54]. The
antioxidant activity is a potential mechanism by
which lycopene may contribute to the prevention of a
variety of oxidative damages, toxicity and other
diseases.
In addition, a reduction in plasma glucose
concentration with stimulation of insulin secretion
that is inconsistent with the result mentioned by El-
Missiry & El-Gindy [55], who reported that the active
principles from plant sources might act by several
mechanisms, such as stimulation of insulin secretion,
increasing the antioxidative capability, enhancing the
effect of insulin and adrenaline as well as increasing
repair/proliferation of pancreatic b-cells confirmed in
our study by gradually increasing the level of insulin
 378 M. M. Ali & F. G. Agha
content in pancreatic tissues with increasing lycopene
doses (Table I).
Kashiwagi et al. [56] reported that elevation of
glucose concentration reduces the activity of GPx,
leading to an accumulation of H2O2. H2O2 catabolism
leads to formation of the superoxide radical anion,
while the decrease in plasma glucose concentration
causes activation of the pentose phosphate pathway,
inactivation of the sorbitol pathway and, consequently,
an increase in NADPH level [57]. NADPH is a co-
factor required for the re-synthesis of reduced glu-
tathione, which regulates the GPx activity and indirect
activity of other antioxidative enzymes [58]. Therefore,
the increased NADPH level may lead to activation of
Scand J Clin Lab Invest Downloaded from informahealthcare.com by Boston University on 07/24/13
all examined antioxidant enzymes, while the decreased
level of hydrogen peroxide and lipid peroxidation
observed after lycopene administration may point to a
reduction in free radical production. Lycopene there-
fore acts as an antidiabetic agent through lowering the
free radical that may be the cause of diabetes.
Consistent with our result, lycopene exhibited a
dose-dependent improvement in the lipid profile,
which is in accordance with Fuhrman et al. [59] and
Agarwal & Rao [60], who mentioned that lycopene
For personal use only.
may have a cholesterol synthesis-inhibiting effect that
enhances LDL degradation. Dietary supplementation
of carotenoids may act as moderate hypocholester-
olaemic agents, secondary to their inhibitory effect on
macrophage 3-hydroxy-3- methyl glutaryl coenzyme
A (HMGCoA) reductase, the rate-limiting enzyme in
cholesterol synthesis. These observations have impli-
cations for both heart disease prevention, through
modification of the processes of cellular atherogenesis
resulting in unstable plaque formation, and the
process of carcinogenesis [61].
There is clear evidence from this study that
oxidative damage plays a major role in STZ-induced
hyperglycaemia, as evidenced by significant inhibi-
tion of the antioxidant defence mechanism accom-
panied by an elevation of hydrogen peroxide and
lipid peroxidation end products. Intraperitoneal
administration of lycopene may reveal their action
on glucose metabolism through reduction of oxida-
tive stress intensity in hyperglycaemia. The protective
effect of lycopene may be connected with the
normalization of hyperglycaemia, the inhibition of
glucose auto-oxidation and, as a result, exhibited a
potent significant amelioration of oxidative stress in
diabetic animals. Also, this study suggests that
lycopene has an improvement effect on serum lipid
profile which reaches the normal level.
Declaration of interest: The authors report no
conflicts of interest. The authors alone are respon-
sible for the content and writing of the paper.
References
[1] American Diabetes Association. Evidence-based nutrition
principles and recommendations for the treatment and
prevention of diabetes and related complications (Position
Statement). Diabetes Care 2003;26Suppl 1:S5161.
[2] Yajnik CS. The insulin resistance epidemic in India: fetal
origins, later lifestyle, or both? Nutritional Rev 2001;59:19.
[3] Banfi C, Eriksson P, Giandomenico G, Mussoni L, Sironi L,
Hamsten A, Tremoli E. Transcriptional regulation of PAI-1
gene by insulin. Diabetes 2001;50:152230.
[4] Barnett P, Braunstein GD. Diabetes mellitus. In: Andreoli TE,
Carpenter CCJ, Griggs RC, Loscalzo J, editors. Cecil
essentials of medicine, 5th edn. Philadelphia: W. B.
Saunders; 2001.
[5] Larner J. Insulin and oral hypoglycemic drug, glucogan. In:
Gilman AG, Goodman LS, Rall IW, Murad F, editors. The
pharmacological basis of therapeutics, 7th edn. NewYork:
Macmillan; 1985. p. 1490516.
[6] The WHO Expert Committee on Diabetes Mellitus. Technical
Report Series World Health Organization, Geneva; 1980.
[7] Opara EC. Oxidative stress, micronutrients, diabetes mellitus
and its complications. J R Soc Health 2002;122:2834.
[8] Abou-Seif MAM, Youssef H. Oxidative stress and male IGF-
1, gonadotropin and related hormones in diabetic patients.
Clin Chem Lab Med 2001;39:61823.
[9] Velazquez E, Winocour PH, Kesteven P, Alberti KG, Laker
MF. Relation of lipid peroxides to macrovascular disease in
type 2 diabetes. Diabet Med 1991;8:7528.
[10] Moncado S, Palmer RMJ, Higgs EA. Nitric oxide: physiology,
pathophysiology and pharmacology. Pharmacol Rev
1991;43:10942.
[11] Abdel-Naim AB, Abdel-Wahab MH, Attia FF. Protective
effects of vitamin E and probucol against gentamicin-induced
nephrotoxicity in rats. Pharmacol Res 1999;40:1837.
[12] McCall MR, Frei B. Can antioxidant vitamins materially
reduce oxidative damage in humans? Free Radic Biol Med
1999;7/8:103453.
[13] Maldonado PD, Barrera D, Medina-Campos ON, Hernandez-
Pando R, Ibarra-Rubio ME, Pedraza-Chaverri J. Aged garlic
extract attenuates gentamicin-induced renal damage and
oxidative stress in rats. Life Sci 2003;73:254356.
[14] Pedraza-Chaverri J, Gonzalez-Orozco AE, Maldonado PD,
Barrera D, Medina-Campos ON, Hernandez-Pando R. Diallyl
disulfide ameliorates gentamicin-induced oxidative stress and
nephropathy in rats. Eur J Pharmacol 2003;473:718.
[15] Cohen LA. A review of animal model studies of tomato
carotenoids, lycopene, and cancer chemoprevention. Exp Biol
Med 2002;227:8648.
[16] Tapiero H, Townsend DM, Tew KD. The role of carotenoids
in the prevention of human pathologies. Biomed
Pharmacother 2004;58:10010.
[17] Gerster H. The potential role of lycopene for human health. J
Am Coll Nutr 1997;16:10926.
[18] Rao AV, Agarwal S. Role of lycopene as antioxidant
carotenoid in the prevention of chronic diseases: a review.
Nutrition Res 1999;19:30523.
[19] Zhang LX, Cooney RV, Bertram JS. Carotenoids enhance gap
junctional communication and inhibit lipid peroxidation in
C3H/10T1/2 cells: relationship to their cancer chemopreven-
tive action. Carcinogenesis 1991;12:210914.
[20] Levy J, Bosin E, Feldman B, Giat Y, Miinster A, Danilenko
M, Sharoni Y. Lycopene is a more potent inhibitor of human
cancer cell proliferation than either alfa-carotene or beta-
carotene. Nutr Cancer 1995;24:25766.

[21] Lingen C, Ernster L, Lindberg O. The promoting effect of
lycopene on the nonspecific resistance of animals. Exp Cell
Res 1959;16:38493.
[22] Bell RH, Hye RJ. Animal models of diabetes mellitus:
physiology and pathology. J Surg Res 1983;35:43360.
[23] Mayhan WG, Simmons LK, Sharpe GM. Mechanism of
impaired responses of cerebral arteriols during diabetes
mellitus. Am J Phys 1991;260:31926.
[24] Diwadkar-Navsariwala V, Novotny Dura J, Gustin GM,
Rodvold KA, Crowell AJ, Sosman JA, Stacewicz-Sapuntzaki
J, Bowen EP. Physiological compartmental model describing
the dynamics of lycopene metabolism in healthy men. J Lipid
Res 2003;44:192739.
[25] Miettinen PJ, Ustinov J, Ormio P, Gao R, Palgi J, Hakonen
E, Juntti-Berggren L, Berggren P, Otonkoski T.
Downregulation of EGF receptor signaling in pancreatic islets
causes diabetes due to impaired postnatal b-cell growth.
Scand J Clin Lab Invest Downloaded from informahealthcare.com by Boston University on 07/24/13
Diabetes 2006;55:3299308.
[26] Wolf SP. Ferrus ion oxidation in presence of ferric ion
indicator xylenol orange for measurement of hydroperoxides.
Meth Enzymol 1994;233:1829.
[27] Conrad CC, Grabowski DT, Walter CA, Sabla S, Richardson
A. Using MT2/2 mice to study metallothionein and oxidative
stress. Free Rad Biol Med 2000;28:44762.
[28] Granger DL, Taintor RR, Boockvar KS. Determination of
nitrate and nitrite in biological samples using bacterial nitrate
reductase coupled with the Griess reaction. Methods
Companion Meth Enzymol 1995;7:7883.
[29] Aebi H. Catalase. Method of enzymatic analysis. New York:
For personal use only.
Academic Press; 1984.
[30] Marklund S, Marklund G. Involvement of superoxide anion
radical in the autooxidation of pyrogallol and a convenient
assay for superoxide dismutase. Eur J Biochem 1974;47:
46974.
[31] Paglia ED, Valentine WN. Studies on the quantitative and
qualitative characterization of erythrocytes glutathione per-
oxidase. J Lab Clin Med 1967;70:15869.
[32] Soto CO, Perez BL, Favari LP, Reyes JL. Prevention of
alloxan-induced diabetes mellitus in the rat by silymarin.
Comp Biochem Physiol 1988;119C:12530.
[33] Jeon SM, Song SH, Jang MK, Kim YH, Nam KT, Jeong TS,
Park YB, Choi MS. Comparison of antioxidant effects of
naringin and probucol in cholesterol-fed rabbits. Clin Chem
Acta 2002;317:18190.
[34] Hunt JV, Smith CC, Wolf SP. Autoxidative glycosylation and
possible involvement of peroxides and free radicals in LDL
modification of glucose. Diabetes 1990;39:14204.
[35] Haluzik M, Nedvidkova J. The role of nitric oxide in the
development of streptozotocin-induced diabetes mellitus:
experimental and clinical implications. Physiol Res 2000;49:
3742.
[36] Tsai EG, Hirsch IB, Brunzell JD, Chait A. Reduced plasma
peroxyl radical trapping capacity and increased susceptibility
of LDL to oxidation in poorly controlled IDD. Diabetes
1994;43:101014.
[37] Baynes JW. Reactive oxygen in the aetiology and complica-
tions of diabetes. In: Ioannides C, Flatt PR, editors. Drug,
diet, and disease. Mechanistic approach to diabetes.
Hertfordshire: Ellis Horwood; 1995.
[38] Evans JL, Goldfine ID, Maddux BA, Grodsky GM. Are
oxidative stress-activated signaling pathways mediators of
insulin resistance and b-cell dysfunction? Diabetes 2003;52:18.
[39] Laight DW, Carrier MJ, Anggard EE. Antioxidants, diabetes,
and endothelial dysfunction. Cardiovasc Res 2000;47:45764.
[40] Piconi L, Quagliara L, Ceriello A. Oxidative stress in diabetes.
Clin Chem Lab Med 2003;41:11449.
Effect of lycopene on hyperglycaemia 379
[41] Escobar JA, Rubio MA, Lissi EA. SOD and catalase
inactivation by singlet oxygen and peroxyl radicals. Free
Rad Biol Med 1996;20:28590.
[42] Blum J, Fridovich I. Inactivation of glutathione peroxidase by
superoxide radical. Arch Biochem Biophys 1985;240:5008.
[43] Kono Y, Fridovich I. Superoxide radical inhibits catalase. J
Biol Chem 1982;257:57514.
[44] Sowers JR. Hyperglycemia-related alterations in vascular nitric
oxide production. J Diabetes Complications 2001;15:614.
[45] Gary A, Grundy SM. Management of dyslipidemia in
NIDDM. Diabetes Care 1990;13:15369.
[46] Levey AS, Beto JA, Coronado BE, Eknoyan G, Foley RN,
Kasiske BL, Klag MJ, Mailloux LU, Manske CL, Meyer KB,
Parfrey PS, Pfeffer MA, Wenger NK, Wilson PW, Wright JT Jr.
Controlling the epidemic of cardiovascular disease in chronic
renal disease: What do we know? What do we need to learn?
Where do we go from here? Am J Kidney Dis 1998;32:853906.
[47] Mulec H, Johnson SA, Bjorck S. Relation between serum
cholesterol and diabetic nephropathy. Lancet 1990;335:
15378.
[48] Laakso M. Glycemic control and the risk of coronary heart
disease in patients with non-insulin-dependent-diabetes-melli-
tus. The Finnish studies. Am J Clin Nutr 1996;124:12730.
[49] Lamarche B, Tchernof A, Moorjani S, Cantin B, Dagenais
GR, Lupien PJ, Despres JP. Small, dense low-density
lipoprotein particles as a predictor of the risk of ischemic
heart disease in men: prospective results from the Quebec
cardiovascular study. Circulation 1997;95:6975.
[50] Britton G. Structure and properties of carotenoids in relation
to function. FASEB J 1995;9:15518.
[51] DiMascio P, Kaiser S, Sies H. Lycopene as the most effective
biological carotenoid singlet oxygen quencher. Arch Biochem
Biophys 1989;274:5328.
[52] Reifen R, Nissenkorn A, Matas Z, Bujanover Y. 5-ASA and
lycopene decrease the oxidative stress and inflammation
induced by iron in rats with colitis. J Gastroenterol 2004;39:
51419.
[53] Stahl W, Sies H. Antioxidant activity of carotenoids. Mol
Aspects Med 2003;24:34551.
[54] Wertz K, Siler U, Goralczyk R. Lycopene: modes of action to
promote prostate health. Arch Biochem Biophys 2004;430:
12734.
[55] El-Missiry MA, El-Gindy AM. Amelioration of alloxan
induced diabetes mellitus and oxidative stress in rats by oil
of Eruca sativa seeds. Ann Nutr Metab 2000;44:97100.
[56] Kashiwagi A, Asahina T, Ikebuchi M, Tanaka Y, Takagi Y,
Nishio Y, Kikkawa R, Shigeta Y. Abnormal glutathione
metabolism and increased cytotoxicity caused by H2O2 in
human umbilical vein endothelial cells cultured in high glucose
medium. Diabetologia 1994;32:2649.
[57] Sinclair AJ. Free radical mechanisms and vascular complica-
tions of diabetes mellitus. Diabetes Rev 1993;2:710.
[58] Togashi H, Shinzawa H, Wakabayashi H, Nakamura T,
Yamada N, Takahashi T, Ishikawa M. Activities of free
oxygen radical scavenger enzymes in human liver. J Hepatol
1990;11:2005.
[59] Fuhrman B, Elis A, Aviram M. Hypocholesterolemic effect of
lycopene and beta-carotene is related to suppression of
cholesterol synthesis and augmentation of LDL receptor
activity in macrophages. Biochem Biophys Res Commun
1997;233:65862.
[60] Agarwal S, Rao AV. Tomato lycopene and low density
lipoprotein oxidation: a human dietary intervention study.
Lipids 1998;33:9814.
[61] Heber D, Yi lu Q. Overview of mechanisms of action of
lycopene. Exp Biol Med 2002;227:9203.