Influence of three medicinal plant extracts on lipid
peroxidation in streptozotocin induced diabetic rats
A P Attanayake, K A P W Jayatilake, C Pathirana, L K B Mudduwa
Faculty of Medicine, University of Ruhuna.
• Traditional knowledge of medicinal plants has always
guided the search for new cures.
• Virtually every indigenous culture in the world uses
medicinal plants in some form or the other for
treatment of ailments.
• A multitude of plant materials have been
described for the treatment of diabetes mellitus
throughout the world.
• The herbal medicine has grown in popularity for
the treatment of diabetes mellitus in Sri Lanka for
a number of reasons.
•
Free radical mediated oxidative stress is a critical
etiological factor implicated in several human diseases
including diabetes mellitus (Bakirel et al., 2008).
• Oxidative stress
Persistent and chronic hyperglycaemia
Depletes the activity of antioxidative defense system
Promotes de novo free radical generation
• Free radicles react with biological molecules
PUFA - most
susceptible
Reaction with these
cell membrane
constituents
Lipid peroxidation
• Any compound possessing radicle scavenging/
antioxidant properties might alleviate the damage
of lipid peroxidation
Use of therapeutic agents
derived from medicinal plants
seems to be an attractive
alternative which could be used
to treat/prevent diabetic
complications induced by
oxidative stress
Medicinal plants selected for the study
bark
leaves
Coccinia grandis
(Cucurbitaceae)
Gmelina arborea
(Verbenaceae)
Extract : aqueous, refluxed
(Jayaweera, 1982)
bark
Spondias pinnata
(Anacardiaceae)
From previous studies
• Efficacy and dose response of the plant extracts on glucose
tolerance in streptozotocin induced diabetic rats
The minimum effective dose of each extract was found
• In vitro antioxidant activity of selected medicinal plant
extracts
possess antioxidant activities with high levels of polyphenolic compounds
The antidiabetic activity and in vivo antioxidant activity of
selected plant extracts
Biochemical parameters
blood/serum
Histopathological
assessment
Kidney, liver
body weight
food intake
water intake
liver
in vivo
antioxidant
activity
Antioxidant markers
and enzymes
Lipid peroxidation
pancreas
Histopathological and
immunohistochemical
assessment
(Haddad et al., 2012)
Objective
• To evaluate the effect of aqueous extracts of Coccinia
grandis, Gmelina arborea, Spondias pinnata on the
extent of lipid peroxidation in streptozotocin induced
diabetic rats.
Diabetic rat model ; streptozotocin (65 mg/kg, ip)
Wistar rats (n=6, b.wt. 200 ±2 5 g)
Rats with fasting blood glucose concentration 180.00 > mg/dL
was considered as hyperglycaemic and used for experiments.
Experimental design
Group
No
Group
1
2
3
Healthy untreated rats
Diabetic untreated rats
Diabetic rats + Coccinia grandis (0.75 mg/kg)
4
Diabetic rats + Gmelina arborea (1.00 mg/kg)
5
Diabetic rats + Spondias pinnata (1.00 mg/kg)
6
Diabetic rats + Glibenclamide (0.50 mg/kg)
*
0
7
14
21
Treatment continued for 30 days
* Collected liver tissue to determine extent of
lipid peroxidation in liver homogenates
28 30
Days
Estimation of the extent of lipid peroxidation
• Malondialdehyde (MDA) is formed as a catabolic product in the lipid
peroxidation
• The common method of measuring MDA is based on the reaction with
thiobarbituric acid (TBA).
TBA + MDA
95 °C for 30 min
MDA-TBA adduct + 2 H2O
pink colored complex
measured the absorbance at 532nm
• MDA values are obtained using the extinction coefficient(ε) 1.56 x 105 M-1 cm-1
•
Results were expressed as nmol/100 mg protein
(Muriel et al., 2001; Okawa et al., 1979)
•
•
•
The results were evaluated by one way ANOVA and
Dunnett’s multiple comparison test.
p<0.05 was considered as statistically significant.
Ethical approval was obtained from the Ethical
Review Committee, Faculty of Medicine, Galle.
Results
Effect of plant extracts on the MDA level in liver homogenates
Concentration of
MDA
(nmol/100mg
protein)
40.00
35.00
30.00
25.00
20.00
15.00
10.00
5.00
0.00
Concentration of MDA
Healthy
untreted
rats
Diabetic
untreted
rats
11.84
36.39
Diabetic rats
Diabetic rats Diabetic rats Diabetic rats
+
+ C. grandis + G. arborea + S. pinnata glibenclami
de
31.10
26.50
28.65
22.10
• The increased levels of malondialdehyde in liver tissue
of streptozotocin induced diabetic rats served as an
index of elevated lipid peroxidation in diabetic
condition.
• The increase in lipid peroxidation indicates increased
oxidative stress, generating more free radicals.
• Significant reduction in lipid peroxidation may be
attributed to the antioxidant activity of the selected
aqueous extracts
Conclusion
• Aqueous leaf extract of Coccinia grandis, bark
extracts of Gmelina arborea and Spondias pinnata
provided protection against lipid peroxidation in
liver tissue in diabetic rats.
References
Dhawan B.N., Srimal R.C. (1998). Acute toxicity and gross effects. In: “Laboratory manual for
pharmacological evaluation of natural products”. United Nations Industrial Development
Organization and International Center for Science and High Technology: 17-20.
Hadded P.S., Musallam L., Martineau L.C., Harris C., Lavoie L. (2012). Comprehensive evidencebased assessment and prioritization of potential antidiabetic medicinal plants: a case study from
canadian eastern james bay cree traditional medicine. Evidence based Complementary and
Alternative Medicine, 10: 1155.
Jayaweera D.M.A. (1982). Medicinal plants used in Ceylon. National Science Foundation of Sri
Lanka, M D Gunasena & Co, Part 1-5.
Muriel P., Alba N., Perez-Alvarez V.M., Shibayama M., Tsutssumi V.K. (2001). Kupfer cells inhibition
prevents hepatic lipid peroxidation and damage induced by carbon tetrachloride. Comparative
Biochemistry and Physiology, 130: 219-226.
Okawa H., Ohrnishi N., Yagi K., (1979). Assay for lipid peroxides in animal tissues by the
thiobarbituric acid reaction . Analytical Biochemistry, 95: 351-358.
Acknowledgement
Financial assistance by UGC/ICD/CRF 2009/2/5.
Mrs B M S Malkanthie and Mr G H J M Priyashantha
Department of Biochemistry, Faculty of Medicine, University
of Ruhuna for laboratory assistance.
Thank you