Ray et al.

RESEARCH ARTICLE

Formulation, Characterisation and

assessment of Antidiabetic activity of

Polyherbal tablet (PHF) in diabetic rat

model

Biswaranjan Ray

, B B Panda

and P K Biswal

Abstract

Diabetes mellitus is a complex and a diverse group of disorders that disturbs the metabolism of carbohydrate, fat and protein.

The number of diabetes mellitus cases has been increasing worldwide in recent years. In 2000, the WHO estimated a total of 171

million of people with diabetes mellitus from the global population, and this report projected to increase to 366 million by 2030.

Now a days polyherbal formulations made by different herbal pharmaceutical company is very much popular and acceptable for

chronic use in case of diabetes, hypertension, bronchial asthma, hyperlipidemia, rheumatoid arthritis etc. As the incidence of

diabetes is increasing day by day globally and the rate of occurrence of disease in India is the high, the popularity of polyherbal

formulation is increasing day by day. Many physicians from different system of medicine refer to advice polyherbal formulations

for long time use in patient of diabetes, as the manufacturer of herbal formulator claim the formulation is having negligible side

effect. Present Study was done with Evaluation of antihyperglycemic Potential of prepared polyherbal Formulation. PHF i.e. with

100mg/kg and 150 mg/kg, the FBS level was reduced from 187.16 ± 3.25 to 133.16 ± 2.93 and 181.5 ± 4.42 to 136.33 ± 2.58,

respectively. In the combination regimen of PHF (150 mg/kg) and Metformin (50mg/kg) the mean FBS level dropped from 180.3

± 1.75 to 131.16 ± 2.14. In all these groups the mean FBS level has reached to the normal pre induction level. The daily single

administration of PHF formulation (150mg/kg) and Metformin (50 mg/kg) signiﬁcantly reduced blood sugar levels of Streptozotocin

(STZ) induced diabetic rats. It is also important to study antihyperglycemic effect of the said Polyhedral Formulation in different

animal Model to conclude exact pattern of Anti hyperglycaemic activity.

Keywords: Diabetes mellitus; polyherbal formulations; antihyperglycemic; Tamarindus indica; S.rebaudiana

Introduction

Diabetes mellitus is a metabolic disease characterized by hyper-

glycaemia. It occurs due to either defects in insulin secretion

or insulin action, or both. The chronic hyperglycaemia is re-

lated with damage and malfunction of different organs, predomi-

nantly kidneys, eyes, nerves, blood vessels and heart. Symptoms

Correspondence: bipinbihari030481@gmail.com

Dept. Pharmacology, College of Pharmaceutical Science, Puri, Odisha,

India

Full list of author information is available at the end of the article.

of marked hyperglycaemia include polyuria, polydipsia, occa-

sionally polyphagia, weight loss and blurred vision. [1]

Over the past 40 years, prevalence of diabetes mellitus has

increased. The trend for future is a continuous increase of all

ethnic groups, men or women, for all age groups worldwide [2].

This increase is observed above all in type 2 diabetes melli-

tus (T2DM) [3]. In 1995, nearly 135 million people were af-

fected and an increase of 300 million cases is estimated for

year 2025 [4]. Various complications of the disease are coronary

artery disease (CAD) including obesity, dyslipidemia, hyperten-

International Journal of Phytomedicine

2021;13(1):005-008

DOI:10.5138/09750185.2461

Received: 13 Dec 2020, Accepted:24 Jan 2021

sion (HT) and physical inactivity [5] . Obesity increases the risk

of CAD in adults and is associated with insulin resistance in nor-

moglycemic as well as in individuals with type 2 DM [6] .

The fruit Tamarind is obtained from the plant Tamarindus in-

dica. The plant is a long lived and large sized tree. It is a famous

as well as common tree found in India. Its fruit is salted and

stored in almost every house. The people of Deccan largely con-

sume it. They say that ‘life is very ticklish in absence of Imli’.

It is also used largely as a ﬂavour, stabilizer and binder in food

preparations. Generally people use Imli to make pickles, curries,

jam and sauce and to prepare majoon as well as jawarish. The

seeds are made ﬂour for making bread in famine seasons. The

seeds are also fried and consumed by poorer. It is very useful for

people of hot area and near to equator.

Tamarind seed powders were used for tannase production by

A. niger. The tannase yield was 6.44 IU/g ds for tamarind seed

powder.

According to Ethnobotanical and traditional literature, seeds

act as anti-asthmatic, antiulcer as well as antioxidant agent.

Vaginal Atony is treated by a pessary of seed kernel. The fried

seed paste was used on anus after setting the tract in proper po-

sition for treatment of rectal prolapse. In Unani system, sper-

matorrhoea, nocturnal emissions as well as seminal debility are

cured by use of roasted kernel and seeds. Homeopathic system

uses seed for the treatment of stomachache. It is also used in

cough and for the relaxation of uvula. The urethral discharge and

polyuria are treated by administration of seed kernel pounded

with milk. The red outer covering of seeds is very useful in di-

arrhea and dysentery. The seeds are used for treatment of colitis

and other intestinal disorders. To feed cattle ground seeds are

usually useful. Seeds are helpful in vaginal discharges and ul-

cers.

Seeds are rich in phenolic compounds, polymeric tannins,

glycosides, fatty acids, ﬂavonoids, saponins, alkaloids. There is

also presence of fatty acids, essential amino acids in seeds. Aux-

ins are available in seeds. Tamarind seeds have 2-hydroxy-3

, 4

dihydroxyacetophenone (TAO), methy l-3, 4-dihydroxybenzoate

(TA1), 3, 4-dihydroxyphenylacetate (TA2) and (-)-epicatechin.

Seed also consist of acetic acid, arabinose, dihydroxylphenyl

acetate. Oil obtained from seeds consists of lauric, palmitic,

myristic, stearic, arachidic, behenic, lignoceric acids, loeic and

linoleic acids. Dry form of kernel has 17.1-20.1% protein, 6.0-

7.4% fat, 65.1-72.2% carbohydrates and some crude ﬁbre and

ash. Whereas roasted kernel includes calcium 121 mg and phos-

phorous 237 mg per 100 gm. It also contains proteins like pro-

lamines and albumins [4].

For hundreds of years, indigenous peoples in Brazil and

Paraguay have used the leaves of stevia as a sweetener. The

Guarani Indians of Paraguay call it kaa jheé and have used it to

sweeten their yerba mate tea for centuries. They have also used

stevia to sweeten other teas and foods and have used it medici-

nally as a cardiotonic, for obesity, hypertension, and heartburn,

and to help lower uric acid levels.

In addition to being a sweetener, stevia is considered (in

Brazilian herbal medicine) to be hypotensive, diuretic, car-

diotonic, and tonic. The leaf is used for obesity, cavities, hyper-

tension, fatigue, depression, sweet cravings, and infections. The

leaf is employed in traditional medical systems in Paraguay for

the same purposes as in Brazil [5–7].

Lufa cylindrical belongs to cucurbitaceae family having king-

dom Plantae according to botanical classiﬁcation. The different

plant parts such as leaves, ﬂower and seeds have been used tradi-

tionally for medicinal purpose. The plant part traditionally used

for intestinal worm, Chronic bronchitis, protozoal disease and

for certain kind of fungal infection [8].

Materials and Method

Plant material and Extraction

Stevia (S.rebaudiana) leaves were collected from palisree mela

(local festival) of Western Odisha from a vendor who cultivate

stevia in small scale for commercial purpose.

Lufa cylindrical belongs to cucurbitaceae family having king-

dom Plantae according to botanical classiﬁcation. The leaves

of the plant were collected from jamadarpli forest area of sam-

balpur district, Odisha.

Stevia leaves were washed to remove dust and subjected to

aqueous extraction after air drying. The dried ground leaves

were mixed with hot water (65 C) at the ratio of 1:45(w/V). The

mixture was shaken and kept at room temperature for 24 hour. It

was sired at least 3-4 times per day. After 24 h,the mixture was

ﬁltered through what man ﬁlter paper and the ﬁlter was evapo-

rated by using vacume evapoarator.

The leaves of lufa cylindrica were dried in shade and pow-

dered to get a coarse powder. About 800gm of dry coarse powder

was extracted with ethanol (40-60

C) by continuous hot perco-

lation using Soxhlet apparatus. The extraction was continued for

72hours. The ethanolic extract was ﬁltered and concentrated to

a dry mass by using vacuum distillation.

Ether and ethanol extracts of fruits were prepared sequen-

tially by standard procedures in soxhalation apparatus. Matured

unripe fruits were shade dried until properly dried, crushed in a

mechanical grinder into ﬁne powder. The powder (500 g) was

extracted sequentially with 1 litres of ether, 1 litres of ethanol in

a Soxhlet apparatus at 65

◦

C until the powder became exhausted

totally. The resulting extracts were ﬁltered, concentrated and

Ray et al. : International Journal of Phytomedicine, 2021;13(1):005-008

006

dried in vacuo (yield 7.60, 8.25 and 8.75% w/w, respectively).

The extracts were stored in desiccators for use in subsequent ex-

periments [9].

Preparation of Polyherbal Tablet

The dried Plant extract of all proposed plants were mixed with

different excipients using wet granulation method for preparing

later solid pharmaceutical form. These prepared granules of each

form were compressed into tablet using compressing machine at

Gayatri College of Pharmacy [10].

Characterisation of Polyhedral Tablet

The Prepared Polyherbal Tablet had been undergone characteri-

sation under following parameter such as particle size measure-

ment, colour and appearance, angle of Repose, Wet variation,

Hardness and Friability test were conducted successfully by tak-

ing standard parameters.

Screening of Antidiabetic Potential of Polyherbal formulation

(PHF)

Animals Sprague Dawley (SD) rats of either sex weighing

between 150-200 gm were procured from Scientiﬁc Trader,

Baleswar. They were housed in polypropylene cages (six rats

/ cage) and maintained under controlled room temperature (20-

◦

C) and with relative humidity of 45-55% under 12:12hr light

and dark cycle. They were provided with standard lab diet and

water ad libitum and kept for 1 week to acclimatize with the

laboratory condition before starting the experiment.

The rats were grouped into 06 groups and each group has 6

rats. Group 1: considered as control group; Group 2: considered

as treatment group, treated with standard antidiabetic compound,

metformin. Group 3, 4 & 5 each were considered as test group,

treated with polyherbal formulation “PHF” in different doses.

Group 6 consist of 06 rats and treated with combination of both

“PHF” and standard drug “metformin” [11–13].

Acute toxicity study

The acute toxicity study was performed as per OECD (guide-

lines 425) and LD

was calculated accordingly. The animals

were examined at every 30 min up to a period of 3 h and then

occasionally for additional 4 h period, ﬁnally 24 h mortality was

recorded. All the animals found to be safe [14].

Experimental induction of Diabetes mellitus

Diabetes mellitus (Type 2 diabetic) was induced in the rats with

normal blood glucose levels by a single injection of streptozo-

tocin (STZ) 35mg/ kg body weight I.P in 0.1M sodium citrate

Table 1 The grouping of rats into different groups for the treatment

of plant extract and the standard drug.

group drug Dose (mg/kg) treatment

I. Normal saline 1ml/kg Control

II. metformin 50.00 Standard

III. PHF 50 Test

IV. PHF 100 Test

V. PHF 150 Test

VI. Metformin +PHF 50+150 Combination

buffer, PH 4.5. In contrast rats in control group were injected

with 0.1M citrate buffer solution only. After injection of STZ

the animals were kept for observation for 48 hrs.

Blood Sample Collection The blood samples were collected

through the tail vein puncturing with a needle. A drop of blood

of size 0.3 to 1.0 µl required. The drop of blood obtained by

pricking with the lancet provided with glucometer for estimation

of FBS and PPBS (Dubois 2014). FBS and PPBS at 1hr and 2hr

were estimated with the help of glucometer weekly for 4 weeks.

The glucometer include a clock that was set for date and time

and memory for past test results. It enables to keep a record of

blood glucose levels over days and wee

Results and Discussion

The mean FBS level of all the rats before induction of diabetics

was found to be 126.5 ± 1.378 mg/dl (Table 1). In contrast the

mean ± S.D of FBS level for different treatment groups over a

time period from week 0 to week 4 was included in Table 2. In

the control group the mean FBS has dropped from 181.16 ± 8.50

in Week 0 to 181.16 ± 2.32. This is far above the mean level

before induction, even after 4 weeks. In Metformin (50mg/kg)

treated group the FBS level was declined from 190.5 ± 2.66 to

138.83 ± 2.23. Similarly in PHF(50mg/kg) treatment group the

FBS was declined from 182.3 ± 6.02 to 135.33 ± 3.01 which

is signiﬁcant and nearer to the pre induction level. In slightly

higher concentration of treatment of PHF i.e. with 100 mg/kg

and 150 mg/kg, the FBS level was reduced from 187.16 ± 3.25

to 133.16 ± 2.93 and 181.5 ± 4.42 to 136.33 ± 2.58, respec-

tively. In the combination regimen of PHF (150mg/kg) and Met-

formin (50mg/kg) the mean FBS level dropped from 180.3 ±

1.75 to 131.16 ± 2.14. In all these groups the mean FBS level

has reached to the normal pre induction level.

One way ANOVA has been applied for comparison among

different treatments groups and followed by Dunnetts multiple

comparison tests for comparison between pair of treatment ef-

fects.

Ray et al. : International Journal of Phytomedicine, 2021;13(1):005-008

007

Table 2 Descriptive statistics of fasting blood sugar at different time period with different doses of drug treatment

Time Period

Mean FBS ± SD (mg/dl)

Control Metformin

50mg/kg

DiaM 50 mg/kg DiaM 100 mg/kg DiaM 150 mg/kg Metformin 50mg/kg

+DiaM 150 mg/kg

Week-0 181.16 ± 8.50 190.5 ± 2.66 182.3 ± 6.02 187.16 ± 3.25 181.5 ± 4.42 180.3 ± 1.75

Week-1 179.13 ± 3.54 144.66 ± 4.50 140.16 ± 1.94 144.5 ± 4.59 141.16 ± 2.86 142.16 ± 3.54

Week-2 179.66 ± 1.75 141.33 ± 2.58 139.33 ± 1.63 139.83 ± 1.94 139.16 ± 1.94 136.16 ± 2.04

Week-3 180.5 ± 1.87 140.5 ± 2.26 133.16 ± 1.83 134.16 ± 1.47 135.16 ± 1.94 133.33 ± 2.66

Week-4 181.16 ± 2.32 138.83 ± 2.23 135.33 ± 3.01 133.16 ± 2.93 136.33 ± 2.58 131.16 ± 2.14

Conclusion

The daily single administration of PHF formulation (150mg/kg)

and Metformin (50 mg/kg) signiﬁcantly reduced blood sugar

levels of Streptozotocin (STZ) induced diabetic mice. This Anti

hyperglycaemic effect is believed to be due to the ability of

PHF extract to stimulate glucose uptake into cells. Coroso-

lic acid, lagerstroemin, tannic acid and penta-O-galloyl-D-

glucopyranose (PGG) are the known active components in

the leaf extract. Metformin (50 mg/kg) did not cause signiﬁcant

glucose reduction. Further study in the ﬁeld of Formulation

needed to overcome manufacturing difﬁculty as well as making

a suitable herbal solid doses form. It also important to study

antihyperglycemic effect of the said Polyhedral Formulation

in different animal Model to conclude exact pattern of Anti

hyperglycaemic activity.

Conﬂict of Interest

There is no conﬂict of interest

Author details

Dept. Pharmacology, College of Pharmaceutical Science, Puri,

Odisha, India.

PG Department, Gayatri College of Pharmacy,

Sambalpur, Odisha, India.

References

[1] Vanmathi SM, Star MM, Jishala MI, Sundaram S. R: A

pathophysiological approach of macrovascular complica-

tion in diabetes mellitus with hypertension- A systemic

review. Research Journal of Pharmacy and technology.

2019;(12):901–906.

[2] Haris MI, Flegal KM, Cowie CC. Prevalence of diabetes,

impaired fasting glucose, and impaired glucose tolerance

in U.S. adults: the Third National Health and Nutrition Ex-

amination Survey. Diabetes Care. 1944;21:518–524.

[3] The wealth of India. Raw materials. 1962;.

[4] Suralkar AA, Rodge KN, Kamble RD, Maske KS. Eval-

uation of anti-inﬂammatory and analgesic activities of

Tamarindus indica seeds. Inter J Pharma Sci Drug Res.

2012;4(3):213–217.

[5] Satyavathi GV, Raina MK, Sharma M. Indian Medicinal

plants. vol. II. New Delhi: ICMR; 1976.

[6] Kujur RS, Singh V, Ram M, Yadava HN, Singh KK, Ku-

mari S, et al. Antidiabetic activity and phytochemical

screening of crude extract of Stevia rebaudiana in alloxan-

induced diabetic rats. Pharm Res. 2010;2(4):258–63.

[7] Abou-Arab AE, Abou-Arab AA, Mf AS. Physico-

chemical assessment of natural sweeteners steviosides pro-

duced from Stevia rebaudianaBertoni plant. Afr J Food

Sci. 2010;4:269–81.

[8] Laidani Y, Hanini S, Henini G. Use of ﬁber Luffacylindrica

for waters treatment charged in copper.Study of the possi-

bility of its regeneration by desorption chemical. Energy

Procedia. 2011;6:381–388.

[9] Harborne JB. Phytochemical methods. 3rd ed. and others,

editor. London: Chapman and Hall; 1998.

[10] Lachman L, Lieberman HA, Kanig JL. The theory and

practice of industrial pharmacy. Lea & Febiger

Philadelphia; 1976.

[11] Hossain A, Sr NU, Salim MDA, Razaul H. Phytochem-

ical and Pharmacological screening of Coccinia grandis

Linn. Journal of Scientiﬁc and Innovative Research.

2014;3(1):65–71.

[12] Knowler WC, Barrett-Connor E, Fowler SE. Diabetes Pre-

vention Program Research Group. Reduction in the inci-

dence of type 2 diabetes with lifestyle intervention or met-

formin. N Engl J Med. 2002;346:393–403.

[13] Maiti R, Das JD, Ghosh UK, D. Antidiabetic ef-

fect of aqueous extract of seed of Tamarindus indica in

streptozotocin-induced diabetic rats. J Ethnopharmacol.

2004;92(1):85–91.

[14] Combes RD, Gaunt I, Balls M. A Scientiﬁc and Ani-

mal Welfare Assessment of the OECD Health Effects Test

Guidelines for the Safety Testing of Chemicals under the

European Union REACH System. ATLA. 2004;32:163–

208.

Ray et al. : International Journal of Phytomedicine, 2021;13(1):005-008

008