This study evaluated the antidiabetic activity of Gluconarc (AYU-DIA), an Ayurvedic formulation, in alloxan-induced diabetic rats. Rats were divided into groups: normal control, diabetic control, groups treated with AYU-DIA doses of 105mg/kg or 210mg/kg, and a group treated with glibenclamide. AYU-DIA at 210mg/kg showed significant reduction in blood glucose in glucose tolerance and adrenaline hyperglycemia tests compared to controls. In alloxan-induced diabetes, AYU-DIA at 210mg/kg and glibenclamide significantly reduced blood glucose over 7 days compared to diabetic controls

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Indo American Journal of Pharmaceutical Research, 2016 ISSN NO: 2231-6876
EVALUATION OF ANTIDIABETIC ACTIVITY OF GLUCONARC (AYU-DIA) AN
AYURVEDIC FORMULATION IN ALLOXAN INDUCED DIABETES MELLITUS IN
RATS
Vijayalaxmi Hiremath1
, Akshatha N1
, Dr. Sunitha R Joshi2
, Dr.Swarna Ranjita2
, Elumalai.P2,
Dr.
Shivalinge Gowda KP*1
1
PES College of Pharmacy, 50 ft Road, Hanumanthanagar, Bengaluru-560050.
2
R & D Team - Vasishta Pharmaceuticals Pvt. Ltd, Banashankari 3rd
stage, Bengaluru-560085.
Corresponding author
Dr.Shivalinge Gowda KP
Asso Prof and HOD,
Pharmacology Department,
PES College of Pharmacy,
50 ft Road, Hanumanthanagar,
Bengaluru-560050
Copy right © 2016 This is an Open Access article distributed under the terms of the Indo American journal of Pharmaceutical
Research, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
ARTICLE INFO ABSTRACT
Article history
Received 19/04/2016
Available online
30/04/2016
Keywords
Alloxan Monohydrate,
Adrenaline,
Glucose Tolerance Test,
Hypoglycaemia.Glucokinase.
The antidiabetic activity of Gluconarc (AYU-DIA) an Ayurvedic formulation was evaluated
its hypoglycemic activity, glucose tolerance test (GTT) in normal rats, adrenaline induced
hyperglycemic rats and alloxan induced diabetes mellitus in albino Wistar rats. 1% CMC
1ml/kg po used as vehicle. In alloxan model rats were divided into five groups of 6 each.
Diabetes was induced for all the groups (II, III, IV, V) except normal control (group I) by
single intraperitoneal administration of alloxan monohydrate in rats (120 mg/kg.ip). AYU-Dia
was administered orally at a dose of 105, 210mg/kg. to the group III and IV. The standard
glibenclamide (4mg/kg po) was administered to the group V. The treatment was made for 7
days. Blood was withdrawn by tail vein puncture and blood glucose was estimated before and
after 24 h the study period. The blood sugar was measured using glucometer and strips. . The
AYU-DIA (105mg/kg.po) not showed hypoglycaemic effect when compared to normal rats.
AYU-DIA (210mg/kg) showed significant decrease in the blood sugar level when compared
to adrenaline induced hyperglycaemic rats. In glucose tolerance test (GTT) AYU-DIA treated
rats showed decreased blood sugar levels. Gluconarc (AYU-DIA 210mg/kg) treated rats
showed significant decrease in the blood sugar level when compared to alloxan induced
diabetic rats. The antidiabetic effect of Gluconarc AYU-DIA is may be due to protection of
inhibition of glucokinase caused by the alloxan in the pancreatic beta cells. It may also due to
prevention of ROS formation in the pancreatic beta cells.
Please cite this article in press as Vijayalaxmi Hiremath et al. Evaluation of antidiabetic activity of AYU-DIA an Ayurvedic
formulation in alloxan induced diabetes mellitus in rats. Indo American Journal of Pharmaceutical Research.2016:6(04).

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INTRODUCTION
Diabetes mellitus is a group of metabolic disorders characterized by altered fuel metabolism, which results in hyperglycemia
and dyslipidemia. It is a serious chronic disease without a cure, and it is associated with significant morbidity and mortality.
According to International Diabetes Federation’s (IDF) 80% of the world diabetic mellitus will be from low- and middle income
countries in 2030. As per IDF 2011 report, China, India and the United States of America have a diabetic population of 90.0, 61.30
and 23.7 million, which may be increased to 129.7, 101.2, and 29.3 millions; respectively1
. The acute complications of diabetes
mellitus are due to severe hyperglycemia. Chronic complications are characterized by damage, dysfunction, and failure of various
organs like the eyes, kidneys, nerves, heart and brain2
.
Insulin is a polypeptide hormone (containing 51 amino acids) secreted by the beta cells of islets of Langerhans of pancreas.
Insulin is packaged within the granules and stored in the β cells, from which it is released at a slow rate by exocytosis into circulation.
When glucose is available, glucose enters the cell via GLUT2 transporters in the plasma membrane and is metabolized to generate
ATP. Intra cellular ATP concentration increases. ATP binds with ATP sensitive K+
channel (an ion channel made up of 8 subunits-
octamer). This inhibits the K+
/ATP channel and K+
conductance decrease. This results in depolarization of the membrane of the beta
cell. This activates the voltage gated Ca2+
channels and thereby stimulates an influx of Ca2+
. Ca2+
mediates the insulin-containing
secretary vesicles within the plasma membrane, leading to insulin secretion.
The insulin receptors are expressed on the energy storing tissues like liver, muscle, and adipose tissue. Binding of insulin to
the extracellular portion of the insulin receptor activates the intracellular tyrosine kinase, resulting in “autophosphorylation” of
tyrosine on the nearby β subunit and in phosphorylation of several other intracellular proteins- insulin receptor-substrate proteins
(IRS-proteins). Phosphorylated IRS proteins interact with many other signalling proteins to effect changes in cellular function,
including glucose transport (via the translocation of GLUT4 glucose transport to the cell surface), protein synthesis and glycogen
synthesis.
In the medical field treating diabetes mellitus without any side effects is still a challenge. In day today practice most of the diabetic
patients were treated with standards anti diabetic drugs such as sulfonylureas and bigunides etc. These drugs have some kind of side
effects like nausea, vomiting, abdominal pain, diarrhoea, head ache etc., and thus search for a new safe and potent anti diabetic herbal
formulation drug is essential to overcome this problems3
.
Traditional medicines all over the world have advocated the use of herbs to treat diabetes mellitus since long time. Many
Indian plants have been studied for their benefits in diabetes mellitus and reports are available in numerous journals. In Ayurvedic
books like Charak, Samhitha, Madhav Nidan, and Astang Sanghra, mentioned the usefulness of about 600 plants having antidiabetic
property. Gluconarc (AYU-DIA) is an indigenous Ayurvedic formulation developed by Vashista Pharmaceutical Pvt Ltd, Bangalore.
This formulation has been selected to study its antidiabetic activity in allaxon induced diabetic rats and probable underlying
mechanism of action of action.
MATERIALS AND METHODS
The Gluconarc (AYU-DIA) sample used for the present investigation was obtained from M/S Vasishta Pharmaceuticals Pvt
Ltd, Bengaluru 560085. Animals- Wistar albino rats (8-10 weeks) (both male and female) were procured from M/S Raghavendra
Enterprises, Bengaluru. The rats were kept in air conditioned animal house of PES College of Pharmacy. The rats were housed in
spacious, hygienic, polypropylene cages with paddy husk as bedding during the experiment period. After randomization into various
groups and before commencement of experiment, the rats were acclimatized for a period of 7 days under standard environmental
conditions of temperature, relative humidity, and dark/light cycle. The animals were fed with water and rat pellet feed (M/s Krish
Scientific Co, Bengaluru) ad libitum. All the animal experimental procedures and protocols used in this study were reviewed by
Institutional Animal Ethics Committee (PESCP/13/IAEC/2015 Dated- 10-1-2015) and were in accordance with the guidelines of the
CPCSEA registration N0- 600/PO/Ere/S/02/CPCSEA).
Methodology-
Study of hypoglycemic activity of AYU-DIA on normal rats.
Normal fasted rats were used for the study of hypoglycaemic activity of AYU-DIA. Overnight fasted rats were used in this
study. Twenty four rats were divided into four groups of six each. Vehicle (1% CMC 1ml/kg po) was administered to the group I.
AYU DIA (105, 210mg/kg po) was administered to the group II and III respectively. Glibenclamide (4mg/kg po) was administered to
the group IV. Blood samples were collected prior and 1, 2, 4 and 6h after the administrating and blood glucose was estimated using
glucometer and strips and the values were expressed in mg/dL. Percentage fall in blood glucose was calculated.
Glucose Tolerance Test (GTT) in normal rats.
The rats were kept for fasting for a period of 15h without food and with sufficient water. Vehicle (1% CMC 1ml/kg po) was
administered to the group I. Group II received only glucose. AYU DIA (105, 210mg/kg po) was administered to the group III and IV
respectively. Glibenclamide (4mg/kg po) was administered to the group V. After 30min glucose 1g/kg was administered by oral
gavage to individual rats of all groups except normal control. Blood samples were collected by tail vein just before the glucose
administration and 2 h after the glucose administration. Blood glucose was measured and expressed in mg/dL.

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Effect of AYU-DIA in normal and adrenaline induced hyperglycemic rats-
The rats were kept for fasting for a period of 15h without food and with sufficient water. Vehicle (1% CMC 1ml/kg po) was
administered to the group I. Group II received only adrenaline. AYU DIA (105, 210mg/kg po) was administered to the group III and
IV respectively. Glibenclamide (4mg/kg po) was administered to the group V. After 1h adrenaline (0,8mg/kg ip) was administered to
all rats group. Blood was withdrawn by tail vein puncture and blood glucose was estimated before and 2h after the adrenaline
administration.
Effect of AYU-DIA in normal and alloxan induced diabetic rats-
Rats were divided into five groups of 6 each. Diabetes was induced for all the groups except normal control (group I) by
single intraperitoneal administration of alloxan monohydrate in rats (120 mg/kg.ip). . Two days after alloxan injection, rats with
plasma glucose levels of 140 mg/dl were included in the study. Treatment with AYU- DIA /glibenclamide started 48 h after alloxan
injection Vehicle (1% CMC 1ml/kg po) was administered to the normal (group I) and diabetic rats (II). AYU DIA (105, 210mg/kg po)
was administered to the group III and IV respectively. Glibenclamide (4mg/kg po) was administered to the group V. The treatment
was made for seven days. Blood was withdrawn by tail vein puncture and blood glucose was estimated before and after the study
period.
RESULTS
Study of hypoglycemic activity of AYU-DIA on normal rats
Table No 1 Showing the hypoglycaemic effects of AYU-DIA in rats.
GP n=6 Treatment Blood sugar level mg/dL
I Normal 115.8+ 1.905
II AYU-DIA (105mg/kg.po) 114.7±2.642
III AYU- DIA (210mg/kg) 107.7±3.084*
IV Glibenclamide 4mg/kg po 88.33 + 5.524 ****
Each value is expressed as Mean ± SEM (n = 6) animals in each group. The comparison was done by One way ANOVA followed by
Sidak's multiple comparisons test.
From the results the group IV rats treated with glibenclamide have shown significant hypoglycaemic effect when compared to the
normal rats. The AYU-DIA (105mg/kg.po) not showed hypoglycaemic effect when compared to normal rats. However rats, treated
with AYU-DIA (105mg/kg.po) had shown little hypoglycaemic effect.
Glucose Tolerance Test (GTT) in normal rats.
Table-2 Effect of AYU DIA on GTT in rats.
Group n=6 Treatment (route-oral) Blood sugar level mg/dL
I Normal control (vehicle treated) 132.7±4.088
II Diabetic control (glucose 1gm/kg) 161.2±14.98
III AYU-DIA (105mg/kg) + glucose 1gm/kg 130.8±7.846
IV AYU-DIA (210mg/kg)+ glucose 1gm/kg 105.8±4.423
V Glibenclamide (4mg/kg) + glucose 1gm/kg 86.83±5.896*
Each value is expressed as Mean ± SEM (n = 6) animals in each group. The second group was compared to the first group by non
paired t test followed by P value 0.0123 as compared to second group with remaining groups by One way ANOVA followed.
The rats administered with glucose (1gm/kg po) had shown increased blood sugar level (161.2±14.98 mg/dL) when
compared to normal rats (132.7±4.088 mg/dL). The rats treated with AYU-DIA (105, 210 mg/kg po) showed decreased blood sugar
levels (130.8±7.846, 105.8±4.423 mg/dL) when compared to group II rats. The glibenclamide treated rats showed significant decrease
in the blood sugar level (86.83±5.896 mg/dL) when compared to group II rats.

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M o d e l II
Bloodglucoselevel
NormalControl
Diabeticcontrol
0
5 0
1 0 0
1 5 0
2 0 0
2 5 0
T re a tm e n t g ro u p
M o d e l I I
Bloodglucoselevel
Diabeticcontrol
AYU-DIA(105mg/kgbw)
AYU-DIA(210mg/kgbw)
Glibenclamide4mg/kgbw
0
5 0
1 0 0
1 5 0
2 0 0
2 5 0
*
T re a tm e n t g ro u p
Fig – 1 Graph showing the effect of AYU-DIA on glucose tolerance test (OGTT).
Effect of AYU-DIA in normal and adrenaline induced hyperglycemic rats
Table No 3 Effect of AYU DIA in adrenaline induced hyperglycemia in rats.
Group n=6 Treatment Blood sugar level mg/dL
I Normal control(vehicle treated) 120.0±3.416
II Diabetic control (Adrenaline 0.8mg/kg ip) 232.7±7.205****
III AYU-DIA(105mg/kg)+ Adrenaline 0.8mg/kg ip 238.2±17.15
IV AYU-DIA(210mg/kg)+Adrenaline 0.8mg/kg ip 116.8±3.807****
V Glibenclamide(4mg/kg)+Adrenaline 0.8mg/kgip 91.17±12.34****
Each value is expressed as Mean ± SEM (n = 6) animals in each group. The second group was compared to the first group by non
paired t test followed by P value P < 0.0001as compared to second group with remaining groups by One way ANOVA followed
Sidak's multiple comparisons test .
Group II rats treated with adrenaline showed significant increase in the blood sugar level (232.7±7.205 mg/dL) when
compared to normal rats (120.0±3.416mg/dL). Group IV rats treated with AYU-DIA (210mg/kg) and adrenaline showed significant
decrease in the blood sugar level (116.8±3.807mg/dL) when compared to group II rats. Group V rats treated with glibenclamide
(4mg/kg po) also showed significant decrease in the blood sugar level (91.17±12.34mg/dL) when compared to group II rats.
M o d e l III
T re a tm e n t g ro u p
Bloodglucoselevel
N
o
rm
a
l
C
o
n
tro
l
D
ia
b
e
tic
c
o
n
tro
l
0
1 0 0
2 0 0
3 0 0
* * * *
M o d e l III
T r e a t m e n t g r o u p
Bloodglucoselevel
D
ia
b
e
tic
c
o
n
tro
l
A
y
u
-
d
ia
(1
0
5
m
g
/k
g
)
A
y
u
-d
ia
(2
1
0
m
g
/k
g
)
G
lib
e
n
c
la
m
id
e
4
m
g
/k
g
0
1 0 0
2 0 0
3 0 0
* * * * * * * *
Fig No 2- Graph showing the effect of AYU-DIA on adrenaline induced hyperglycemic rats.

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Effect of AYU-DIA in normal and alloxan induced diabetic rats
Table No 4- Effect of AYU DIA in allaxon induced diabetic rats.
Group n=6 Treatment Blood sugar level mg/dL
I Normal control(vehicle treated) 127.0±6.663
II Diabetic control (Alloxan 120mg/kg ip) 229.2±7.736***
III Ayu-dia(105mg/kg)+ Alloxan 120mg/kg ip 191.2±24.84
IV Ayu- dia (210mg/kg)+ Alloxan 120mg/kgip 129.5±7.689***
V Glibenclamide(4mg/kg)+ Alloxan 120mg/kg 118.2±10.68****
Each value is expressed as Mean ± SEM (n = 6) animals in each group. The second group was compared to the first group by non
paired t test followed by P ˂0.0001 compared to second group with remaining groups by One way ANOVA followed Sidak's multiple
comparisons test .
Group II rats treated with alloxan showed significant increase in the blood sugar level (229.2±7.736mg/dL) when compared
to normal rats (127.0±6.663mg/dL). ). Group IV rats treated with AYU-DIA (210mg/kg) and alloxan showed significant decrease in
the blood sugar level (129.5±7.689 mg/dL) when compared to group II rats. Group V rats treated with glibenclamide (4mg/kg po) also
showed significant decrease in the blood sugar level (118.2±10.68 mg/dL) when compared to group II rats.
M o d e l IV
T re a tm e n t g ro u p
Bloodglucoselevel
N
o
rm
a
l
C
o
n
tro
l
D
ia
b
e
tic
c
o
n
tro
l
0
1 0 0
2 0 0
3 0 0
** *
M o d e l I V
T r e a t m e n t g r o u p
Bloodglucoselevel
Diabeticcontrol
Ayudia(105mg/kg)
Ayudia(210mg/kg)
Glibenclamide4mg/kg.
0
1 0 0
2 0 0
3 0 0
* * * * * * *
* * *
Fig No 3- Graph showing the effect of AYU- DIA on alloxan induced diabetic rats.
DISCUSSION
From the results the group IV rats treated with glibenclamide showed significant hypoglycaemic effect when compared to the
normal rats. This hypoglycaemic effect of glibenclamide is due its stimulatory effect on insulin secretion. The AYU-DIA
(105mg/kg.po) not showed hypoglycaemic effect when compared to normal rats. However rats, treated with AYU-DIA (105mg/kg.po)
had shown little hypoglycaemic effect. The two metabolites of glibenclamide 4-trans-hydroxy-glibenclamide and 3-cis-hydroxy-
glibenclamide are responsible for the hypoglycaemic effect. These metabolites stimulate the secretion of insulin from beta cells of the
pancreas4
. The hypoglycaemic effect of AYU-DIA may be due to its stimulatory effect on beta cells of the pancreas.
The oral glucose tolerance test (OGTT) measures the body’s ability to use glucose, the main source of energy. In OGTT the
glucose is given and blood samples taken afterwards to determine to quickly it is cleared from the blood. Elevated blood sugar levels
observed in diabetes, acromegaly. The rats administered with glucose (1gm/kg po) had shown increased blood sugar level when
compared to normal rats. The rats treated with AYU-DIA showed decreased blood sugar levels when compared to group II rats. The
glibenclamide treated rats showed significant decrease in the blood sugar level when compared to group II rats.
Adrenaline increases blood sugar level by transient increase in hepatic glucose production and an inhibition of glucose
disposal by insulin dependent tissues. Adrenaline augments hepatic glucose production by stimulating glycogenolysis and
gluconeogenesis5
. The reduction of blood sugar level caused by ATU-DIA may be due to blockage of adrenaline induced
hyperglycemia.
Alloxan inhibits the glucose induced insulin secretion in the beta cells of the pancreas. This increases the blood sugar level.
This is due to selective inhibition of glucokinase enzyme in the beta cells. Through the release of reactive oxygen species (ROS)
alloxan also causes the selective necrosis of pancreatic beta cells. The molecular shape of alloxan is similar to glucose. Alloxan and
glucose molecules are hydrophilic in nature. Hence they cannot pass through the cell membrane. The allaxon enter the beta cells
through glucose transporter (GLUT II). Inhibition of glucokinase reduces glucose oxidation and ATP generation, thereby suppressing
the ATP signal that triggers insulin secretion. Inhibition of glucokinase is achieved within 1 min of exposure to alloxan6
.

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Group II rats treated with alloxan showed significant increase in the blood sugar level when compared to normal rats. Group
IV rats treated with AYU-DIA and alloxan showed significant decrease in the blood sugar level when compared to group II rats.
Group V rats treated with glibenclamide also showed significant decrease in the blood sugar level when compared to group II rats. The
antidiabetic effect of AYU-DIA is may be due to protection of inhibition of glucokinase caused by the alloxan in the pancreatic beta
cells. It may also due to prevention of ROS formation in the pancreatic beta cells.
CONCLUSION
From the results of the present study, it is concluded that Gluconarc (AYU-DIA) an Ayurvedic formulation developed by M/s
Vasishta Pharmaceuticals Pvt. Ltd, Bengaluru, possesses antidiabetic properties that may serve to protect users against Type II
diabetes mellitus. Further research could be done to determine its specific mechanism of action.
Conflict of interest
The authors hereby declare no conflict of interest.
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2. Pittas AG and Greenberg AS. Contemporary diagnosis and management of diabetes. Handbooks in Health Care Co. Newtown,
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3. Anbu N, Musthafa MD, Velpandian V. Anti-diabetic activity of polyherbal formulation Aavaraiyathi churnam in alloxan induced
diabetic rats. International Journal of Toxicological and Pharmacological Research. 2013; 4(4), 77-80.
4. Rydberg T, Jonsson A, Roder M, Melander A. Hypoglycemic activity of glyburide (glibenclamide) metabolites in humans.
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6. Lenzen S. Mechanisms of allaxon and streptozotocin induced diabetes. Diabetelogia. 2008;51:216-26.
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