Phytochemical Studies on Selected Medicinal Plant Gymnema Sylvestre

Phytochemical studies on selected medicinal plant Gymnema sylvestre.

Keywords:

Phytochemical, Anti-inflammatory, Antimicrobial, TLC, Physicochemical analysis, Fluorescent analysis.

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Dates: Received: 06 Apr 2012 Accepted: 19 Apr 2012 Published: 21 May 2012

Article Citation: Gnana Sangeetha D and Jegadeesan M. Phytochemical studies on selected medicinal plant Gymnema sylvestre. Journal of Research in Plant Sciences (2012) 1: 077-082

Original Research

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Authors:

Gnana Sangeetha D1.

and Jegadeesan M2.

Institution:

1. Department of Chemistry,

PSNA College of

Engineering and Technology,

Dindigul, Tamil Nadu, India

2. Department of

Environmental and Herbal

Sciences, Tamil University,

Thanjavur, Tamil Nadu,

India.

Corresponding author:

Gnana Sangeetha D.

Web Address: http://plantsciences.info/

documents/PS0027.pdf.

Journal of Research in

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ABSTRACT: Gymnema sylvestre R. (Br).(Asclepiadaceae) is a common medicinal plant available in Tamil Nadu is a woody, climbing plant that grows in the tropical forest of central and Southern India. It also happens to be a first-rate warrior against diabetes. The plant is called Gud-mar (Gud-Jaggery, mar-kills) in India and well known for masking sweet taste. It is reported to be effective against many chronic diseases is screened for its phytochemical content, microbial activity and anti-inflammatory activity. Extracts (Alcoholic, aqueous, acetone and hexane) from the plant is prepared and analyzed. Qualitative phytochemical tests are done to detect the presence of Carbohydrate, Alkaloid, Tannins, Phenols, Saponins, Fixed oils, Gums and Mucilage. Quantitative methods like phytochemical analysis, gravimetric estimation, RBC membrane stabilization and TLC profiles are used to determine the active principle Gymnemic acid, anti-inflammatory activity and antimicrobial activity. The results showed that the leaf extracts studied contain the bio active compounds phenols, alkaloids, tannins, saponins and Gymnemic acid. Water extract of normal var. had positive reaction for carbohydrate (Molisch reagent) hairy var. leaves had negative reaction. The presence of active phytochemical substances with antimicrobial and anti-inflammatory activities may provide substantial basis for the use of this plant in ethno medicine.

Journal of Research in Plant Sciences An International Scientific Research Journal

INTRODUCTION:

India has been the rich source of plant based

drugs. Most of the Indian populations especially in rural

areas use traditional medicines only. On realizing the fact

that herbal medicine has no side effect (Bhandari and

Grover 1998, Shetty and Sunderasan, 1997) and many

more herbal medicines are reported to be effective

against many chronic disease for which there were no

allopathic drugs, production of herbal based medicine

has increased tremendously throughout the world. In

modern India extracts of Gymnema leaves are used

successfully, either alone or in combination with other

Ayurvedic compounds as a remedy against diabetes

mellitus. The term “Destroyer of sugar” is tradionally

used for Gymnema because chewing the leaves will

abolish the taste of sweetness. That is sweet food no

longer tasted sweet, but rather became almost completely

tasteless. The medicinally active parts of the plant are the

leaves and roots. Gymnema sylvestre is an Indian

medicinal plant used for diabetes in folk medicine,

Siddha and Ayurveda. Modern scientific studies also

proved its efficacy against the disease (Gent et al., 1999).

Gymnema sylvestre leaf extracts decreased the blood

sugar level in varying degrees. Blood sugar lowering unit

(BLU) of activity of each leaf extract

(Chatopadhyay;1999).Amrycard composed of Gymnema

sylvestre was tried in 20 patients of diabetes for three

weeks.Dose was determined on the basis of condition of

patients as 3 caps to insulin dependend case,2caps for

moderate case and 1 cap to mild diabetic case.It is

concluded that the drug provides a total beneficial

therepy in all type of diabetes and tends to increase

insulin secretion from islets of pancreas.(Bhagwan

singh;1997) Gymnema sylvestre is a liane.Two varities

exist in this species;one with normal and the other with

hairy leaf.Leaf in normal variety is lanceolate to ovate

apex acute and base truncate, less pubescent on both

sides.The hairy variant is ovate to cordate in shape, apex

acute and base cordate. With the above background the

present study investigates the pharmacological,

phytochemical properties and anti-inflammatory,

antimicrobial activities of the leaves of normal and hairy

variants of the medicinal plant Gymnema sylvestre.

MATERIALS AND METHODS:

Collection of plant material and identification:

The plant specimen Gymnema sylvestre were

collected from herbal garden, Tamil University,

Tanjavur. Collected plants were carefully examined and

identified with the help of Local Floras. Identities of

specimens are further confirmed with reference to

herbarium sheets at Rapinath Herbarium,

Trichy.Voucher specimen were deposited at Tamil

University, Herbarium.

Extraction:

The leaves of Variants Gymnema sylvestre are

shade dried and mechanically powdered after keeping it

in an oven at 40˚C for 24 hours. These powdered

materials are used for further physiochemical,

phytochemical and fluorescent analysis.

Preliminary phytochemical analysis:

Preliminary phytochemical analysis is done using

the procedures of Kokate (1994), (Balakumar etal; 1998).

Alkaloids, Carbohydrate, Tannins, Phenols, and

Saponins, Fixed oils, gums, mucilage and fats were

quantitatively analyzed. The intensity of the coloration

determines the abundance of the compound present.

Antimicrobial studies:

The aqueous extract of leaves Variants of

Gymnema sylvestre (Alagesaboopathi C. 2011) is used

throughout the study. The aqueous extract 10%, 20% and

40% are tested against different bacterial pathogens

(Microbial Type Culture Collection-MTCC) such as

Staphylococcus epidermis (MTCC-435), Klebsiella

pneumonia (MTCC-2653), Serratia marcens (MTCC-

2645), and Citrobacter diversus for its antimicrobial

activity. It was demonstrated by well diffusion assay.

Sangeetha and Jegadeesan, 2012

078 Journal of Research in Plant Sciences (2012) 1: 077-082

Physicochemical analysis:

Leaves of Gymnema sylvestre are analyzed for

ash value, loss on drying, extractive values, solubility

etc...In different solvents and extracts, using the

procedures recommended in Indian pharmacopoeia

(Anonymous, 1996; 1985; 1966). The colour of the leaf

powder of normal variety is dark green and that of hairy

variant is light green.

Fluorescent analysis: Fluorescent analyses of

leaves of Variants of Gymnema sylvestre in different

chemical reagents were analyzed using both Visible and

UV light.

TLC Profiles:

For TLC separation the alcoholic extracts of

leaves of Variants of selected medicinal plant Gymnema

sylvestre is spotted on silica gel-G coated plates.BAW,

Water, Ferosal and 60%Acetic acid was used as mobile

phase. Equal proportion of iodine chamber was used as

detecting reagent.

RESULTS AND DISCUSSION:

Physiochemical Analysis:

Physiochemical values of leaf of normal, hairy

var. and stored leaf are given in Table 5. It is interesting

to note that all the physiochemical values of normal leaf

and stored leaf differ greatly the values are also differ

between leaves of normal and hairy var. Total ash

content value of hairy var. is grater than that of normal

var. and among normal var. total ash content of fresh

leaves is less (13.7%) than stored leaves. Acid insoluble

ash value for normal var. is less (0.36%) than that for

hairy leaf (0.65%) in normal variant stored leaf

registered more acid insoluble ash value (0.41%). Water

soluble ash content of normal var. was far grater

(39.82%) than that of hairy var. (27.09%) and this value

in stored leaf was also lower than fresh leaves. Sulphated

ash value of normal var. was less (10.43%) when

compare to hairy var. (11.26%) and stored leaf (15.48%).

Normal var. had lower loss on drying. Solubility of leaf

of normal var. is more (29.96%) than that of hairy var.

(20.16%) and stored leaf (28.74%). Extractive value %

of normal var. is more (32.34%) compared to hairy and

stored leaf.

Qualitative phytochemical Analysis:

Leaf extract of Gymnema var. in water and

alcohol positively for all the phytochemical compounds

(Table 1). Though both water and alcohol extracts of

normal and hairy var. had similar phytochemical


Journal of Research in Plant Sciences (2012) 1: 077-082 079

Table -1 Prelimnary phytochemical analysis of gymnema sylvestre

S. No. Compound

Tested Reagent

Water Alcohol

Normal Normal

(stored) Hairy Normal

Normal

(stored) Hairy

1 Carbohydrate Fehling’s - - - - - -

Molish ++ ++ - + ++ ++

2 Alkaloid

Picric +++ +++ ++ ++ +++ +++

Mayer ++ ++ + + ++ ++

Wagner’s +++ +++ +++ +++ ++ -

3 Tannis & Phenols

Fecl3 +++ +++ ++ ++ - -

Gelatin + + ++ ++ ++ ++

Lead acetate +++ +++ +++ +++ ++ -

4 Saponin Foam +++ +++ +++ ++ +++ +++

5 Gum & Mucilage - ++ ++ ++ ++ ++ ++

6 Fixed & Oils - + - + ++ ++ ++

(+++) - Appreciable Amount

(++) - Moderate

(+) - Small Amount

(-) - Completely Absent

constituents, some diagnostic characteristics were noted.

Water extract of normal var. had positive reaction for

carbohydrate (Molish reagent) hairy var. leaves had

negative reaction. Alcohol extracts of normal, stored and

hairy leaves answered positive for carbohydrates. It is to

be noted that water and alcohol extracts of the three leaf

samples had not answered for Fehling’s reagent

indicating absences of reducing sugar. It was also

observed that stored leaf had not answered for oil and fat.

Anti Inflammatory:

The in-vitro anti-inflammatory study (RBC

membrane stabilization) using different concentrations

(10, 25, 50, 75, 100 & 200 µg/ml) of Gymnema aqueous

extracts was carried out. It was found that leaf extract of

normal var. at a concentration of 200 µg/ml showed

membrane stabilization activity with maximum activity

200 µg/ml concentrations (Graph 1&2). It was found that

leaf extract of hairy var. also showed membrane

stabilization activity concentration 200 µg/ml.

Antimicrobial activity:

Water extract of G. sylvestre normal and hairy

var. were screened for antibacterial and antifungal

activity against Staphylococcus epidermis, Klebsiella

pneumonia, Serratia marcens, Citrobacter diversus,

Aspergillus niger and Aspergillus flavus (Table 2).

Antimicrobial activity of aqueous extracts of both

variants revealed that all concentration inhibited growth

of S.epidermis with increase in zone of inhibitions with

increasing concentration. Leaf extract of normal

G.sylvestre had inhibitory activity against K.pneumonia

and A.niger whereas that of hairy var. had no activity

against these organisms. Both variant had activity against

S.marcens whereas hairy var. had inhibitory action

against C.diversus and A.flavus. Normal var. had no

activity.

TLC Studies:

Rf values of aqueous extracts of G. sylvestre leaf

(normal, hairy and stored) were determined on TLC

(Table 4). On TLC when 60% acetic acid was used as

mobile phase, all the three leaf samples had only one

spot Rf value 0.95. In BAW solvent phase, the normal

variant had two spots with Rf values 0.18 and 0.88.

Hairy var. had two spots with Rf values at 0.18 and 0.53.

Stored leaf had only spot corresponding with Rf value

0.16. TLC profile using Ferosal as mobile phase gave a

single spot with Rf value 0.74 for normal leaf, two spots

at Rf values 0.74 and 0.81 for hairy var. and two spots at

0.74 and 0.83 for stored leaf. In water, normal leaf, hairy

leaf and stored leaf exhibited only one spot with Rf value

0.79. Thus the three leaf samples of G.sylvestre had

distinguished.TLC profiles in the four mobile phase

used. Thus the three leaf samples had different profiles,

though they had common spots with similar Rf values.

Thus, as 60% acetic acid as solvent base, all the leaf

samples had only one spot with Rf value 0.95. In BAW,

the entire sample had a band at Rf 0.18. In Ferosal at Rf

0.74 all samples had spot. And in water, all the samples

had only one spot with Rf value at 0.79. This indicates

that while similar biological active compounds are

present in all the samples they differ by either presence

or absence of additional compounds.



Table -2 Antimicrobial activity of aqueous extracts of gymnema sylvestre

S. No Organism

Zone of Inhibition (cm)

Concentration

10% 20% 30%

Normal Hairy Normal Hairy Normal Hairy

1 Staphylococcus epidermis 1.3 0.7 1.5 2.7 2.7 3.0

2 Klebsiella pneumonia 1.0 Nil 1.1 Nil 1.4 Nil

3 Serratia marcesens 1.0 0.9 1.1 1.9 2.1 1.5

4 Citrobactor diversus Nil 0.6 Nil 1.4 Nil 1.7

5 Aspergillus niger 1.4 Nil 2.0 0.8 2.2 1.1

6 Aspergillus flavus Nil 1.2 Nil 1.8 Nil 1.9

Leaves of normal and hairy variants of

Gymnema sylvestre varieties in water and alcohol had

answered positively for all the phytochemical

compounds (Table-1).

Anti-microbial activity (Table-2) of water

extracts of normal and hairy varieties of G.sylvestre was

screened. Both varieties had shown dosage dependent

inhibitory activity against S.epidermis .Normal varieties

had activity against K.pneumoniae,A.niger while hairy

variant had no activity. Hairy variant had action against

C.diversus and A.flavus. Thus two variants of

G.sylvestre have similar and dissimilar antimicrobial

activity against specific organisms.

Physicochemical, organoleptic, quantitative,

microscopical analysis of two varieties of G.sylvestre

revealed both similar and dissimilar characters which can

be used for diagnostic purpose.

Fluorescent analyses of leaf powder of normal

and hairy varities of G.sylvestre in different chemical

reagents. (Table-3) showed no major distinguishing

features for both Visible and UV light. This indicates

that both varieties have similar chemical constituents

Water extract of normal and hairy varieties

exhibited RBC membrane stabilizing activity (Siddiqui,

2000) at 200µg/ml concentration. Thus both varieties

have anti-inflammatory activity (Graph-1&2).


Journal of Research in Plant Sciences (2012) 1: 077-082 081

Table -3 Fluorescent analysis of dried leaf powder of gymnema sylvestre

S. No Powder + Reagent Normal Var. Hairy Var.

Visible UV Visible UV

1 Powder Light Green Light Green Light Green Light Green

2 P + IN HCL Dark Green Dark Green Dark Green Dark Green

3 P + H2SO4 Dark Brown Dark Brown Dark Brown Dark Brown

4 P + HNO3 Yellow Light Yellow Yellow Dark Yellow

5 P + NH4OH Dark Green Dark Green Dark Green Dark Green

6 P+Picric Acid Light Green Light Green Light Green Light Green

7 P+Iodine Light Brown Dark Brown Brown Brown

8 P+FeCl3 Black Brown Black Brown

Table -4 Rƒ values of aqueous extract of gymnema sylvestre leaf

Mobile Phase

Rƒ Value of Gymnema Sylvestre

Normal Hairy Normal (Stored)

Spot-I Spot-II Spot-I Spot-II Spot-I Spot-II

Acetic Acid 0.95 - 0.95 - 0.95 -

BAW 0.18 0.88 0.18 0.53 0.18 -

Ferosal 0.74 - 0.74 0.81 0.74 0.83

Water 0.79 - 0.79 - 0.79 -

Graph 1: Effect of aqueous extract of Gymnema

Sylvestre (Normal) on RBC Lysis

OD

AT

56

0 m

m

Graph 2: Effect of aqueous extract of Gymnema

Sylvestre (Hairy) on RBC lysis

OD

AT

56

0 m

m



TLC profiles of aqueous extracts (Table-4) of

normal, hairy and stored leaves samples of G. sylvestre

revealed diagnostic features. All the three leaf samples

developed only one spot in acetic acid and water,

whereas in BAW and ferosol three samples developed

different bands with different Rƒ values. The TLC study

is a further proof that these two variants are

phytochemically different.

REFERENCES

Alagesaboopathi C. 2011. Antimicrobial screening of

selected medicinal plants Tamilnadu, India. Afr J

Microbiol Res., 56:617-621.

Balakumar T and selvakumar. 1998. Prelimnary

observation on the micro propagation of the antidiabetic

herb Gymnema sylvestre.National Plant Research.A

(52):2-4.

Bhandari V and Grover JK. 1998. A reappraised of

clinical studies on the comparative influence of three

indigenous plant drug in diabetes melliyus.Hamard

m e d i c u s . 4 1 ( 2 ) ; 9 - 1 5 . E n v i b b s i , 2 0 0 4 . h t t p : / /

www.envibbsi.org/medi:htm#Gymnema sylvestre.

Debruyne D. 1997. Clinical pharmacokinetics of

fluconazole in superficial and systemic mycoses. Clin

Pharmacokinet., 33:52-77.

Gent JF, Persand,Hettinger TP, Frank ME and

marks CE. 1999. Tast confusion following gymnemic

acid rinse,chem. Senses.24:393-403.

Shetty BR and Sunderasan. 1997. Studies on the

resayana effect of a geriatric formulation in apparently

normal aged persons.Journal of Research Ayurveda and

Siddha.18 (3-4):108-117.

Siddiqui. 2000. Development in the chemistry and

pharmacology of G.sylvestre.Journal of Medicinal and

aromatic Plant sciences.22. (2-4):222-231.

S.No Parameter Normal Normal

(Stored) Hairy

1. Total Ash 13.71 15.50 17.88

2. Acid-insoluble

Ash 0.36 0.41 0.65

3. Water-soluble

Ash 39.82 18.00 27.09

4. Sulphate Ash 10.43 15.48 11.26

5. Loss on

Drying 3.23 - 4.44

6.

Solubility

Water 29.96 28.74 20.16

Alcohol 10.26 36.02 22.52

7. Extractive

Value Water 32.34 26.62 22.89

8. Gymnemic

Acid 2.75 3.7 3.75

Table-5 Physiochemical Values (Precentage) of

Leaf Powder Gymnema Sylvestre

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Phytochemical Studies on Selected Medicinal Plant Gymnema Sylvestre