Indian Journal of Experimental Biology
Vol. 44, September 2006, pp. 745-748
Biochemical effects of Nigella sativa L seeds in diabetic rats
M Kaleem, D Kirmani, M Asifa, Q Ahmedb & Bilqees Bano∗
Department of Biochemistry, Faculty of Life Sciences, Aligarh Muslim University, Aligarh 202 002, India
a
Department of Medicine, J.N. Medical College, Aligarh Muslim University, Aligarh 202 002, India
b
Department of Pharmaceutical Chemistry, IIUM, Kuantan 25200 Pahang DM, Malaysia
Received 16 March 2005; revised 19 May 2006
Oral administration of ethanol extract of N. sativa seeds (300 mg/kg body weight/day) to streptozotocin induced
diabetic rats for 30 days significantly reduced the elevated levels of blood glucose, lipids, plasma insulin and improved
altered levels of lipid peroxidation products (TBARS and hydroperoxides) and antioxidant enzymes like catalase,
superoxide dismutase, reduced glutathione and glutathione peroxidase in liver and kidney. The results confirm the
antidiabetic activity of N. sativa seeds extract and suggest that because of its antioxidant effects its administration may be
useful in controlling the diabetic complications in experimental diabetic rats.
Keywords: Antioxidants, Diabetes mellitus, Free radicals, Nigella sativa, Oxidative stress
Diabetes mellitus (DM) affects a large number of contain several active constituents such as
people throughout the world and more so in India. thymoquinone, many monoterpenes such as p-cymene
Clinically, the disease is associated with a number of and α-pinene. Kanter et al.12 have reported
chronic complications including nephropathy, antidiabetic activity of N. sativa on streptozotocin
neuropathy, retinopathy and cardiovascular diseases1. induced diabetic rats. The present study has been
In view of undesirable side effects of synthetic drugs2, aimed to investigate the effect of ethanolic extract of
WHO has recommended evaluation of plants effective N. sativa on lipid peroxides and enzymic antioxidants
in different diseases. Many Indian medicinal plants in liver and kidney tissues of streptozotocin (STZ)
have been found to be useful in successfully induced diabetic rats.
managing diabetes and from some of them active
Materials and Methods
principles have been isolated3. Thus, it will be useful
Chemicals—Bovine serum albumin (BSA),
to look for new and if possible more efficacious drugs
thiobarbituric acid (TBA) and streptozotocin were
and the vast reserves of phytotherapy may be an ideal
procured from Sigma Chemical Co., St. Louis, MO,
target. The beneficial effects of some synthetic and
USA. All other chemicals used were of analytical
plant drugs have been shown to be due to their
grade and purchased locally.
antioxidant properties. Plants often contain substantial
Plant material—The N. sativa seeds were collected
amounts of antioxidants, flavonoids and tannins4.
from The Survey of Medicinal Plant Unit, Regional
Implication of oxidative stress in the pathogenesis of
Research Institute of Unani Medicine, Aligarh (U.P)
diabetes is suggested not only by oxygen free-radical
India. Identification of the samples was further
generation but also due to non-enzymatic protein
confirmed with the Department of Botany, Aligrah
glycosylation5 and alteration in antioxidant enzymes6.
Muslim University (AMU), Aligarh and a voucher
Among medicinal plants, Nigella sativa L
specimen (JK 021) was deposited in the department
(Ranunculaceae; called as Kalongi, in Hindi) seeds
herbarium.
have bronchodilator7, antibacterial8, diuretic,
Preparation of plant extract—The N. sativa seeds
hypotensive and immunopotentiating properties10. El-
9
were dried at room temperature. The seeds were
Tahir et al.11 have reported that N. sativa seeds
powdered in an electrical grinder and stored at 5°C
____________ until further use. Seed powder (300 g) was extracted
*Correspondent author
Phone: 91-571-2700741
with petroleum ether (60°-80°C) to remove lipids. It
Fax: 91-571-2702758 was then filtered and the residue was extracted with
E-mail: bilqeesbano@redilfmail.com 95% ethanol by soxhlet extraction. Ethanol was
746 INDIAN J EXP BIOL, SEPTEMBER 2006
evaporated in a rotary evaporator at 40°-50°C under measurements were carried out in a Camspec UV-
reduced pressure. The yield of extract was 15 g and Visible (Camspec M330B, UK) spectrophotometer.
the substance was dissolved in water before use. Statistical analysis—All the data were statistically
Animals—Male albino Wistar rats (32), weighing evaluated with SPSS/7.5 Software. Hypothesis testing
about 150-180 g, obtained from Central Animal methods included one way analysis of variance
House, J.N. Medical College, AMU, Aligarh were (ANOVA) followed by least significant differences
used. The animals were maintained on standard rat test. P values <0.05 were considered statistically
feed supplied by Amruth, Pune. The experiments significant.
were conducted according to the ethical norms
Results
approved by Institutional Animal Ethics Committee
Body and organ weight—A significant (P< 0.05)
(IAEC) Guidelines.
decrease in body (27.3%) and liver weights (9.5%)
Experimental induction of diabetes—Diabetes was
and increase in kidney weight (25%) was observed in
induced using streptozotocin (55 mg/kg body weight
diabetic rats as compared to control rats.
single i.p. injection) in 0.1 M citrate buffer pH
Administration of N. sativa extract to diabetic rats
4.5(ref.13).
increased body (33.5%) and liver (41.3%) weight and
Experimental design—The rats were divided into
decrease kidney (15.7%) weight to near normal level.
following four groups of eight each:
In the group 3 animals treated with insulin, body and
Group 1: Healthy control rats receiving 0.1 M
kidney weights returned to normal values, but liver
citrate buffer (pH 4.5).
weight returned to only 18.4% of normal value.
Group 2: Diabetic controls.
Blood parameters—The levels of blood glucose
Group 3: Diabetic rats treated with protamine-zinc
and plasma lipid profile (TC, LDL-C and TG) were
insulin ip injection (6 units/kg/day) for 30 days.
significantly higher (P< 0.05) in diabetic rats as
Group 4: Diabetic rats given ethanol extract of N.
compared to control rats. Oral administration of N.
sativa (300 mg/kg/day) in aqueous solution orally
sativa extract to diabetic rats, brought down blood
once daily for 30 days.
glucose to near normal and showed significant
Rats in all the groups were provided with food and decrease (P< 0.05) in the level of plasma lipid profile
water ad libitum. The blood glucose and body weight and an increase in plasma insulin level as compared to
were checked weekly. At the end of the experimental untreated diabetic rats (Table 1). The effect was
period, the rats were anaesthetized and sacrificed. however slightly less than that in animals treated with
Blood sample was collected in tubes containing insulin.
potassium oxalate and sodium fluoride. The plasma Lipid peroxidation products—There was a
was stored at -4oC until analysis was completed. The significant elevation in the level of TBARS and
liver and kidney tissues were excised, rinsed in ice- hydroperoxides in both liver and kidney tissues of
cold saline, cut into small pieces and homogenized untreated diabetic rats. Administration of N. sativa
with Potter-Elvehjem glass-Teflon homogenizer in
Table 1―Effect of treatment of diabetic rats with N. sativa seeds
Tris-HCL buffer (pH 7.4). The homogenate was extract for 30 days on their blood glucose, plasma lipid profile and
centrifuged at 10000 rpm for 10 min. Supernatant was insulin levels
used for various measurements. The following [Values are mean ± SD from 8 animals in each group]
analyses were carried out: blood glucose, plasma total Parameter Control Diabetic Diabetic + Diabetic+N.
cholesterol (TC), high density lipoprotein cholesterol Insulin sativa
(HDL-C) and triglycerides (TG) were estimated using TC* 120.6±11.3 283.5±12.5 157.2±10.1a 171.2±6.0a
standard kits of Ranbaxy Laboratories, New Delhi. LDL-C* 62.4±5.54 186.18±7.1 71.4±8.8 85.0±11.1a
Plasma insulin level was assayed by enzyme linked HDL-C* 40.6±4.1 59.8±3.4 68.1±5.4 a
63.5±5.8a
immunosorbent assay (ELISA) kit (Boerhringer LDL/HDL 1.53±0.12 3.11±0.34 1.04±0.45 1.33±0.22
Mannheim, Germany). Low density lipoprotein TG* 89.3±6.7 185.1±12.1 102.4±10.7a 113.8±10.1a
Blood 95.0±8.0 283±15.2 98±7.0a 102±7.3a
(LDL-C)14, thiobarbituric acid reactive substances
Glucose*
(lipid peroxides)15 and hydroperoxides16, reduced Plasma 15.13±0.71 4.22±0.26 10.21±0.47 11.37±0.72a
glutathione (GSH)17, superoxide dismutase (SOD)18, Insulin†
glutathione peroxidase (GPx)19, catalase (CAT)20 and a
P< 0.05 when compared with diabetic rats
protein21, were also estimated. All spectrophotometric Units: *mg/100ml serum; †µUnits/ml serum
KALEEM et al.: BIOCHEMICAL EFFECTS OF NIGELLA SATIVA SEEDS IN DIABETIC RATS 747
extract to diabetic rats significantly decreased considerably, serum lipids of diabetic rats which were
(P< 0.05) the level of TBARS and hydroperoxides however not completely normal. Oxidative stress
(Table 2). The effect of N. sativa extract was slightly plays a role in the causation of diabetes and
more than that with insulin except in kidney in which antioxidants have been shown to have a role in the
the effect of both was equal. alleviation of diabetes24. In diabetes oxygen free
Antioxidants—Significant decreases was observed radicals (OFRs) are generated by stimulating H2O2 in
in the activities of SOD, CAT, GPx and GSH in liver vitro and in vivo in pancreatic β-cells. OFRs
and kidney tissues of untreated diabetic rats. scavenging enzymes normally respond to conditions
Administration of N. sativa seeds extract and insulin of oxidative stress with a compensatory mechanism
considerably improved the activities of these enzymes that increases the antioxidative enzyme activity in
which were however not normal in treated diabetic diabetic rats initially but reduced in chronic
rats (Table 2). uncontrolled diabetes25,26.
The results of the present study indicate
Discussion significantly increased lipid peroxidation of rats
Treatment of diabetic rats with N. sativa seeds exposed to STZ and its statistically significant
extract showed improvement in many parameters. attenuation by N. sativa seeds extract treatment. This
Their body, liver and kidney weights and blood suggests protective role of N. sativa seeds extract
glucose were near normal, indicating possible which could be due to the antioxidative effect of
prevention of muscle wasting known to occur in flavonoids present in the seeds which act as strong
diabetes. The studies confirm the earlier findings that superoxide radicals and singlet oxygen quenchers27.
N. sativa seeds extract has antidiabetic effect12. As The decreased activities of SOD, GPx and CAT in
seen in the present studies also the level of serum both liver and kidney during diabetes may be due to
lipids was usually raised in diabetes and such an production of reactive oxygen free radicals. Treatment
elevation represents a risk factor for coronary heart with N. sativa seeds extract increased the activity of
disease22. Lowering of serum lipid levels through diet these enzymes and thus may help to counteract the
or by drug therapy seems to be associated with a damage by the free radicals generated during diabetes.
decrease in the risk of vascular disease23. Reduced glutathione is a potent free radicals
Administration of N. sativa seeds extract improved, scavenger. Treatment of N. sativa seeds extract
Table 2—Effect of treatment of diabetic rats with N. sativa seeds extract
resulted in the elevation of the GSH levels, which is
for 30 days on superoxide dismutase, catalase, glutathione peroxide, present in the islet β-cells and protects the membranes
reduced glutathione, TBARS and hydroperoxides in liver and kidney against oxidative damage by regulating the redox
[Values are mean ± SD from 8 animals in each group] status of protein in the membrane28.
Groups Control Diabetic Diabetic+Insulin Diabetic+ The present results suggest that it is worthwhile
N. sativa carrying out further studies to find out whether N.
Liver sativa seeds extract could be a supplement, as an
SODa
antioxidant therapy and may be beneficial for
20.16±1.60 14.26±1.30 18.72±2.00* 18.34±3.06*
CATb 0.223±0.030 0.137±0.019 0.181±0.026* 0.210±0.017* correcting the hyperglycemia and preventing diabetic
GPxa 0.187±0.020 0.131±0.014 0.168±0.038* 0.170±0.026* complications due to lipid peroxidation and free
GSHc 55.6±3.00 30.3±2.34 54.4±3.20* 52.7±2.30* radical oxidation. Longer duration studies of N. sativa
TBARSd 0.87±0.05 1.69±0.31 1.01±0.21* 0.93±0.06*
Hydro- 62.5±2.5 88.3±4.1 67.1±3.5* 63.1±1.7*
seeds and its isolated active compounds on chronic
peroxidese models are necessary to develop a potent antidiabetic
Kidney drug from N. sativa seeds.
SODa 12.27±1.48 8.24±0.83 13.15±1.02* 12.18±2.25*
CATb 0.116±0.014 0.072±0.009 0.104±0.018* 0.110±0.012* Acknowledgement
GPxa 0.050±0.009 0.036±0.005 0.048±0.008* 0.046±0.004* The authors are thankful to CSIR and UGC for
GSHc 31.4±1.50 19.2±1.16 26.5±1.95* 30.1±2.00* financial assistance and Dr. S Zehra, S P Baba and
TBARSd 1.25±0.16 2.08±0.22 1.66±0.15* 1.47±0.14*
Hydro- 48.4±3.4 71.8±2.1 59.5±0.9* 60.2±1.4*
M Priyadarshini for help.
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