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Alleviation of alloxan-induced dia-
betes and its complications in rats 
by Actinodaphne hookeri leaf extract 

%-3 



Introduction 

Diabetes mellitus leads to metabolic disorder and 
is characterized by hyperglycemia resulting from 
defects in insulin secretion, insulin action or both 
(Atkinson and Maclaren, 1994). It is also estimat-
ed that there are 30 to 33 million diabetic patients 
in India now and every fourth diabetes patient in 
the world today is an Indian. Indians are genet-
ically more susceptible to diabetes and the World 
Health Organization predicts the number of dia-
betic persons in India would go up to 40 million 

by 2010 and to 74 million by 2025 (Pillai, 2006). 
However, insulin administration is effective to 
some extent in alleviating diabetes mellitus and 
increasing the life expectancy of diabetic patients, 
but there is limitations as well as draw backs for 
this therapy. Some oral hypoglycemic agents are 
also employed in this regard, but they are also not 
without adverse effects.  

Actinodaphne hookeri Meissn. (Family Lauraceae; 
local name: Pisa) is a moderate evergreen tree 
found in many parts of India, more common in 

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Alleviation of alloxan-induced diabetes and its complications in 
rats by Actinodaphne hookeri leaf extract 

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Received:                            28 June 2008 
Accepted:                            10 July 2008 
Available Online:               18 July 2008  
 
DOI: 10.3329/bjp.v3i2.946 
 
 
 
 
 
 
 
 

 
Cite this article: 
Prajapati DD, Patel NM, Savadi RV, 
Akki KS, Mruthunjaya K. Alleviation 
of alloxan-induced diabetes and its 
complications in rats by Actinodaphne 
hookeri leaf extract. Bangladesh J Phar-
macol. 2008; 3: 102-06. 

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Leaves of Actinodaphne hookeri Meissn (Family Lauraceae; local name: 
Pisa) has been in use traditionally for the treatment of diabetes and 
disorders of the urinary tract which are more common in Chattisgarh 
and eastern part of India. In the present study, leaves of A. hookeri 
were subjected to phytochemical investigation and evaluated for anti-
diabetic activity. The ethanol and the chloroform extract were found 
to have significant (p<0.01) blood glucose lowering effect. The extracts 
also significantly (p<0.01) lowered the increased serum cholesterol 
and low density lipoprotein levels. Preliminary phytochemical inves-
tigation revealed the presence of alkaloids, flavonoids, triterpenoids 
and glycosides as the major constituents in the ethanol extract. The 
chloroform extract also showed significant (p<0.01) antihyperglyce-
mic activity and contained alkaloids and triterpenes. It is concluded 
that the anti-diabetic activity of A. hookeri may be due to the presence 
of alkaloids and triterpenes, and might be promising for the develop-
ment of phytomedicine for diabetes mellitus along with its associated 
complications. 

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the Chhattisgarh state and in the eastern part of 
India. Traditionally the leaves are used in the 
treatment of diabetes mellitus and in urinary dis-
orders (Anonymous, 2003, Kirtikar and Basu, 
1995). The phytoconstituents reported are querce-
tin-3-rhamnoside, amorphous alkaloid, hentri-
acontanone and hentriacontanol (Chopra et al., 
1956; Yoganarasimhan, 2000). The present study 
aimed at investigating into the effect of ethanol, 
petroleum ether, chloroform and methanol ex-
tracts of A. hookeri leaves on blood glucose levels 
in normoglycemic and alloxan-induced diabetic 
rats.  

 

Materials and Methods 

Collection of plant material 

A. hookeri leaves were collected in the month of 
June 2006, from Jog falls near a town called Sirsi 
of Karnataka State, India. The plant was authenti-
cated by Dr. B.D. Huddar, Department of Botany, 
H.S. Kotambri Science Institute, Hubli, Karnataka 
State, India. The voucher specimen (No: 05 PG 
358) was deposited in herbarium of Department 
of Pharmacognosy and Phytochemistry, KLES’ 
College of Pharmacy, Hubli, India. 

Preparation of extracts 

Shade-dried and powdered leaves of A. hookeri 
was soxhlet extracted with ethanol (95%). Anoth-
er batch was also soxhlet extracted successively 
with petroleum ether (40-60ºC), chloroform, and 
with methanol in increasing order of polarity. The 
extracts were concentrated under reduced pres-
sure using rotary evaporator and the residue was 
dried in desiccators over anhydrous calcium chlo-
ride.  

Qualitative chemical tests 

All the extracts were tested to know the different 
constituents present in them by the standard pro-
cedures. The extracts were tested for sterols 
(Finar, 1975), alkaloids, triterpenes, saponins 
(Cromwell et al., 1955; Kokate et al., 1996), flavo-
noids (Geinssman et al., 1955), tannins (Trease 
and Evans, 1989), and carbohydrates (Hawks, 
1971). 

Chemicals 

Tween 80 of analytical grade was purchased from 
Hi-Media Labs, India, Glucometer (Accuchek-

Sensor) was purchased from Roche Diagnostics, 
Mumbai, India. Alloxan monohydrate was pur-
chased from Spectrchem Pvt. Ltd Company, 
Mumbai. Glibenclamide from Aventis Pharma 
Ltd., India. All the solvents used for extraction 
were purchased from Ranbaxy Fine Chemicals 
Ltd., New Delhi, India. 

Animals 

All the experiments on animal were conducted 
according to protocols that were approved by the 
Institutional Animal Ethics Committee/
IAEC.Clear/2005-06/I). Anti-diabetic activity was 
carried out using 3 months old healthy albino 
Wister rats of either sex weighing between 150-
200 g. Animals were allowed free access to tap 
water and pellet diet throughout the study. Acute 
toxicity studies were carried out using albino 
mice. 

Acute toxicity study 

Albino mice of either sex weighing between 20-30 
g were used. The animals were fasted over night. 
Acute toxicity study was performed according to 
OECD guidelines; method followed is according 
to number 420. It was found that tolerated dose 
was higher than 5,000 mg/kg body weight 
(OECD, 2001).  

Glucose tolerance test 

Glucose tolerance test was carried out using the 
method described elsewhere (Babu et al, 2002). 
Four groups of 6 rats each were used for the 
study. Group I served as normal control (Vehicle: 
1% Tween 80 emulsion), Group II animals were 
administered with alcohol extract, Group III ani-
mals were administered with petroleum ether 
extract, Group IV animals were administered 
with chloroform extract, Group V animals were 
administered with methanol extract. All the ex-
tracts were administered through oral route at a 
dose of 500 mg/kg. The rats of all the groups 
were administered with 60% glucose (3 g/kg 
orally) 30 min after the administration of extract. 
Blood samples were collected from the tail prior 
to drug administration and at 30, 90 and 150 min 
after glucose administration. Blood glucose levels 
were measured using Glucometer (Accuchek-
Sensor). 

Alloxan-induced diabetes 

The albino rats weighing 150-200 g of either sex 

 Bangladesh J Pharmacol 2008; 3: 102-106                                                               103 



were allowed to fast for 24 hours prior to experi-
mentation and rendered diabetic by a single dose 
of intraperitoneal injection of alloxan 150 mg/kg 
body weight (Vogel and Gang, 2002) After 18 
hours of injection of alloxan, diabetes was con-
firmed by testing urine and blood sugar level 
more than 200 mg/dL were selected for the fur-
ther study. Animals were maintained for four 
days in diabetic condition for well establishment 
of diabetes. They were divided into seven groups. 
Group I: Healthy normal animals received only 
the vehicle (1% Tween 80); Group II: Untreated 
but diabetes-induced animals served as a nega-
tive control; Group III: Diabetes-induced animals 
and treated with standard drug glibenclamide 10 
mg/kg body wt./day orally; Group IV: Diabetic 
animals and treated with ethanol extract; Group 
V: Diabetic animals and treated with petroleum 
ether extract; Group VI: Diabetic animals and 
treated with chloroform extract; Group VII: Dia-
betic animals and treated with methanol extract. 
All the extracts were given orally at the dose of 
500 mg/kg body weight.  After 15 days animals 
were sacrificed under ether anesthesia and blood 
was collected from the carotid artery for the de-
termination of blood glucose level, serum total 
cholesterol, total triglyceride, high density lipo-
protein (HDL), low density lipoprotein (LDL) and 
very low density lipoprotein (VLDL) levels 
(Kannur et al., 2006). 

Statistical analysis 

Comparison between control and drug treated 
groups were analyzed by software Prism 4.03 
with one-way ANOVA. p<0.05 and p<0.01 were 
considered to be significant. 

Results and Discussion 

As shown in Table I, all the groups showed sig-
nificantly (p<0.05) lowered the blood glucose lev-
el compared to those of Group I (treated with glu-
cose only) at 150 min interval. But as the differ-
ence between the blood glucose levels in Group I 
from 30 min interval to 150 min interval was not 
significant. The values at 150 min were not of 
much importance. The reduction in blood glucose 
level which was due to normal metabolism of 
glucose and feed back mechanism (Rang et al., 
2001). So, the increase in blood glucose had auto-
matically reduced at 150 min time interval in case 
of all the extracts. But the Group II (treated with 
ethanol extract) maintained the blood glucose 
level significantly (p<0.01) lower than that of nor-
mal control (Group I), i.e. from 30 to 150 min 
whereas the other groups did not. Group III 
(petroleum ether) lowered the blood glucose level 
significantly (p<0.05) at 30 min but not at 90 min. 
Group IV (chloroform) lowered the blood glucose 
level significantly (p<0.05) at both 30 and 90 min. 
Group V (methanol) lowered the blood glucose 
level significantly (p<0.01) at 30 min and (p<0.05) 
90 min. These results showed that all the extracts 
have hypoglycemic effect at specific time interval. 
Among the four extracts hypoglycemic effect was 
observed more consistently in case of the alcohol 
extract. This would suggests the presence of hy-
poglycemic components in ethanol extract. Chlo-
roform extract contains triterpenes and alkaloids, 
methanol extract contains mainly phenolic 
(flavonoids and tannins) and carbohydrates (data 
not shown). Petroleum ether contains triterpenes 

104 Bangladesh J Pharmacol 2008; 3: 102-106                                                              

Table I 

Effect of various extracts of A. hookeri on blood glucose tolerance in normal rats 

Groups   

0 min 30 min 90 min 150 min 

Glucose only 87.3 ± 3.7 120.0 ± 10.1 105.8 ± 5.2 99.6 ± 4.3 

Glucose + Ethanol extract 78.7 ± 2.4 83.5 ± 0.3 70.8 ± 5.3a 66.0 ± 8.0 

Glucose + Petroleum ether extract 85.7 ± 7.4 111.3 ± 10.4 95.3  ± 7.3a 75.8 ± 3.6 

Glucose + Chloroform extract 87.2 ± 4.3 116.5 ± 16.4 85.0 ± 6.3a 70.0 ± 5.1 

Glucose + Methanol extract 80.3 ± 7.1 90.3 ± 5.8 75.2 ± 4.3b 70.2 ± 8.7 

The values are expressed as mean ± SEM; ap<0.01 when compared to control;  bp>0.05 when compared to control  

Blood glucose level (mg/dL)  



present within it was identified by qualitative 
chemical test and thin layer chromatography. 

As shown in Table II, all the treated groups ex-
cept Group VII significantly (p<0.01) lowered the 
blood glucose level, compared to Group II diabet-
ic control. Group VII failed to lower the increased 
blood glucose level significantly. Methanol ex-
tract contained only carbohydrates and phenolics 
where as petroleum extract contained triterpenes 
and chloroform extract contained both alkaloids 
and triterpenes. The ethanol extract contained all 
the constituents. This indicates that the lowering 
of the blood glucose levels were due to the pres-
ence of triterpenes and alkaloids.  

As shown in the Table III, in the Group II 
(untreated but diabetic-induced) total cholesterol, 
triglyceride, LDL and VLDL level were increased 
significantly (p<0.01) when compared to Group I. 
But HDL level was decreased significantly 
(p<0.01) when compared to Group I. Group III 
and IV significantly (p<0.01) reduced the total 
cholesterol, triglyceride, LDL and VLDL, and sig-

nificantly (p<0.01) increased the HDL level when 
compared to diabetic control (Group II). But these 
values (total cholesterol, triglyceride, LDL, VLDL 
and HDL) were not significant (p>0.05) when 
compared to Group I. Group V decreased the 
VLDL (p<0.01) and LDL (p<0.05) level signifi-
cantly, Group V also decreased the total choles-
terol and triglyceride not significantly, increased 
the HDL significantly. Group VI significantly re-
duced the total cholesterol, triglyceride and 
VLDL but not LDL. But the HDL level was in-
creased close to the normal control. Group VII 
reduced the total cholesterol and VLDL signifi-
cantly when compared to Group II. But not re-
duced the triglyceride and LDL value and also 
failed to increase the HDL value close to normal. 

The effect of ethanol extract of A. hookeri 
(containing triterpenes, alkaloids, flavonoids, tan-
nins and carbohydrates) on lipid parameters ap-
peared better than the other extracts followed by  
chloroform (containing triterpenes and alkaloids) 
and petroleum ether (containing triterpenes). The 
alleviating effect upon lipid parameters appeared 
minimum in case of methanol extract (containing 
carbohydrates and tannins). This appeared simi-
lar to those as observed in case of blood glucose 
level. So, it indicates that the anti-diabetic effect of 
A. hookeri was perhaps due to triterpenes and al-
kaloids. 

 

Acknowledgements 

The authors are grate thankful to Dr. B.D. Huddar, 
Department of Botany, H.S. Kotambri Science Institute, 
Hubli, Karnataka, India, for authenticating the plant 
material. We would like to thank Dr. Purshotam Red-
dy, Pathologist Sadhana Diagnostics, Hubli, India for 

 Bangladesh J Pharmacol 2008; 3: 102-106                                                               105 

Table III 

Effect of Actinodaphne hookeri leaves extracts on lipid profile in diabetic  rats  

Parameters 

Gro u p I  Gro u p I I  Gro u p I I I  Gro u p I V 

Total cholesterol 47.3 ± 3.1 76.3 ± 2.9 51.8 ± 2.5a 53.2 ± 4.1a 

Triglyceride 81.3 ± 2.9 137.3 ± 4.9 93.5 ± 6.3a 108.8 ± 4.8a 

HDL 46.0 ± 2.0 29.7 ± 2.2 47.7 ± 2.5a 46.7 ± 2.3a 

LDL 8.5 ± 0.4 18.7 ± 1.3 10.0 ± 1.3a 10.0 ± 1.5a 

VLDL 8.8 ± 0.8 22.3 ± 1.5 13.3 ± 1.5a 10.5 ± 0.9a 

The values are expressed as mean ± SEM; ap<0.01 when compared to diabetic control; bp>0.05 when compared to diabetic control; cp<0.05 when 
compared to diabetic control 

Blood glucose level (mg/dL) 

Gro u p V  Gro u p VI  Gro u p VI I  

71.3 ± 3.9b 52.3 ± 2.9a 56.7 ± 4.7a 

123.2 ± 7.0b 106.5 ± 5.1a 120.3 ± 5.1b 

39.2 ± 1.9a 46.5 ± 2.7a 31 ± 2.3b 

11.8 ± 1.2c 20.0 ± 2.8b 23.0 ± 2.1b 

9.7 ± 1.2a 8.4 ± 0.8a 10.3 ± 1.3a 

Table II 

Effect of various extracts of Actinodaphne hookeri 
on blood glucose level in rats 

Group Blood glucose level (mg/dL) 

I 72.3 ± 2.4 

II 487.3 ±  6.9 

III 175.3 ± 5.9a 

IV 208.7 ± 6.9a 

V 374.5 ± 16.5a 

VI 222.7 ± 13.7a 

VII 466 ± 7.4b 
The values are expressed as mean ± SEM; ap<0.01 when compared 
to diabetic control; bp>0.05 when compared to diabetic control  



his help in carrying out the biochemical investigations. 

 

References 

Anonymous. The wealth of India: Raw material. In: 
Actinodaphne. New Delhi, Publication and Infor-
mation Directorate CSIR, 2003, pp 70-71. 

Atkinson MA, Maclaren NK. The pathogenesis of insu-
lin dependent diabetes mellitus. New Engl J Med. 
1994; 331: 1428–36.  

Babu V, Gangadevi T, Subramanian A. Anti-
hyperglycemic activity of cassia kleinii leaf extracts in 
glucose fed normal rats and alloxan-induced diabetic 
rats. Indian J Pharmacol. 2002; 34: 409-15.  

Chopra RN, Nayar SL, Chopra IC. Glossary of Indian 
medicinal plants. In: Actinodaphne hookeri. New Delhi, 
Publication & Information Directorate CSIR Press, 
1956, p 6. 

Cromwell BT, Peach K, Tracey MV. Modern methods 
of plant analysis. In: Alkaloids. Vol. 3. 1st ed. Berlin, 
Springer Verlag, 1955, pp 373-74. 

Finar IL. Organic chemistry stereochemistry and chem-
istry of natural products. Vol. 2. London, Longman 
Scientific and Technical Press, 1975, p 518. 

Geinssman TA. Modern methods of plant analysis. In: 
Flavonoids. Vol. 3. Berlin, Springer Verlag, 1955, pp 
467-74. 

Hawks PB. Physiological chemistry. In: Carbohydrates. 
New Delhi, Tata McGraw Hill Publishing Company 
Ltd, 1971, p 373. 

Kannur DM, Hukkeri VI, Akki KS. Anti-diabetic activi-
ty of Caesalpinia bonduucella seed extracts in rats. 
Fitoterapia 2006; 77: 546-49.  

Kirtikar KR, Basu BD. Indian medicinal plants. In: Ac-
tinodaphne. Vol. 3. 2nd ed. Dehradun, India Interna-
tional Book Distributors, 1995, pp 2156-57. 

Kokate CK, Purohit AP, Gokhale SB. Pharmacognosy. 
In: Alkaloid. 26th ed. Pune, Nirali Prakashan, 1996, p 
447. 

OECD. Organization for economic co-operation and 
development. In: Acute oral toxicity guidelines 420, 
Paris, 2001, pp 12-14. 

Pillai M. Reader’s Digest. In: Could you be a diabetic. 
New Delhi, Living Media India Limited Press, 2006, 
p 138. 

Rang HP, Dale MM, Ritter JM, Moore PK. Pharmacolo-
gy. In: The endocrine pancreas and the control of 
blood glucose. London, Churchil Livingstone, 2001, 
pp 385-92. 

Trease GE, Evans WC. Pharmacognosy. In: Phenols 
and phenolic glycosides. London, ELBS, 1989, pp 223
-24, 246-49. 

Vogel GH, Gang W. Drug discovery and evaluation 
pharmacological assay. In: Methods to induce experi-
mental diabetes mellitus. Heidelberg, Springer Ver-
lag, 2002, p 950.  

Yoganarasimhan SN. Medicinal plants of India. In: 
Actinodaphne hookeri. Vol. 1. Bangalore, Interline Pub-
lishing Pvt. Ltd, 2000, p 13. 

 

$XWKRU�,QIR 
Dharmeshkumar D. Prajapati (Principal contact) 
e-mail: dharmesh09mpc @yahoo.co.in 

106 Bangladesh J Pharmacol 2008; 3: 102-106