Turkish Journal of Botany

Volume 34 Number 3 Article 7

1-1-2010

Comparative pharmacognostic studies on the barks of four Ficus

species
K BABU

GOKUL SHANKAR SABESAN

SADANANDA RAI

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Recommended Citation
BABU, K; SABESAN, GOKUL SHANKAR; and RAI, SADANANDA (2010) "Comparative pharmacognostic
studies on the barks of four Ficus species," Turkish Journal of Botany: Vol. 34: No. 3, Article 7.
https://doi.org/10.3906/bot-0907-115
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 Turk J Bot
34 (2010) 215-224
© TÜBİTAK
Research Article doi:10.3906/bot-0907-115

Comparative pharmacognostic studies on the barks of four

Ficus species

Koilpillai BABU1,*, Sabesan GOKUL SHANKAR2, Sadananda RAI1

1
R & D Center, Cholayil Private Limited, 31-A/24, SIDCO Industrial Estate (North Phase), Ambattur, Chennai - 600 098,
Tamilnadu, INDIA
2Microbiology Unit, Faculty of Medicine, AIMST University, Kedah, MALAYSIA

Received: 20.07.2009
Accepted: 04.04.2010

Abstract: The barks of 4 Ficus species, namely F. racemosa, F. virens, F. religiosa and F. benghalensis, are important
ingredients in many Ayurvedic and traditional formulations. The barks are considered to be very effective in various
treatments, such as diabetes, skin diseases, ulcers, and nervous disorders. During market research, we observed that
various species of Ficus barks were sold in Indian market under traditional names, such as Plaksah, Udumbarah,
Asvatthah, and Vatah. The barks of the species mention above are usually interchanged or adulterated with other species
of Ficus because of the limited knowledge in identification and differentiation. Therefore, a detailed comparative
pharmacognostic evaluation of the 4 species has been carried out with the aim to establish the diagnostic keys of these
important drugs based on the macroscopic, microscopic, and HPTLC profiles. Detailed diagnostic and distinctive
characteristics for the differentiation of the 4 Ficus species are discussed.

Key words: Ficus racemosa, Ficus virens, Ficus religiosa, Ficus benghalensis, Moraceae, Bark, Pharmacognosy

Introduction of selecting incorrect raw drugs/adulterants.

Plants are utilized extensively as raw drugs for Therefore, an extensive anatomical and
many formulations in traditional systems of medicine. phytochemical screening is needed for each raw drug
To check the genuineness of the raw drugs and to used in the formulation to avoid any ambiguity and
detect adulteration of these materials, an authentic such a study will serve also as a reference for further
pharmacognostic study is needed for each raw drug. studies (Vaibhav & Kamlesh, 2007). Anatomical
Usually the drugs are collected by traditional studies are helpful in describing a particular drug with
practitioners who have inherited Ayurvedic or other a special emphasize on quantitative microscopy, such
herbal practices. Their identification is mostly based as sclereids, starch grains, crystals, stomata, and
on morphological features or other traditionally trichomes, and qualitative microscopy, such as xylem,
known characteristics. In such cases, there is a chance phloem, and other tissues (Brinda et al., 2000).

* E-mail: [email protected]; [email protected]

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Comparative pharmacognostic studies on the barks of four Ficus species

The genus Ficus belonging to the family Moraceae Phytochemical analysis

constitutes an important group of trees with immense For the chemical analysis all the barks were shade
medicinal value. Among the many species, the most dried separately for 10 days and powdered using a
important are the 4 trees with milky latex, namely Ficus
pulveriser. Powdered samples were subjected to
racemosa L. (Sanskrit - Udumbarah), Ficus virens Aiton
physico-chemical analysis, such as the percentage of
(Sanskrit - Plaksah), Ficus religiosa L. (Sanskrit -
water and alcohol soluble extractive, total ash, acid-
Asvatthah), and Ficus benghalensis L. (Sanskrit -
insoluble ash (Ayurvedic Pharmacopoeia of India,
Nyagrodhah, Vatah), that constitute the group
2001) and preliminary phytochemical screening was
“Nalpamaram” in Ayurveda. The barks of these species
form an important ingredient in many Ayurvedic carried out using standard procedures (Evans, 1989;
formulations, such as Nalpamaradi tailam, Sofowara, 1993; Harborne, 1998).
Chandanasavam, and Saribadyasavam (Sivarajan & HPTLC analysis
Balachandran, 1994). They are used separately or in Two grams of powdered material was refluxed
combination in different formulations. The barks are with 10 mL of methanol in a water-bath at 60 °C for 30
used for various purposes: as an astringent medicine, min, consecutively 3 times, and then concentrated
for cooling in action, as haemostatic, as laxative, in
and dried. The final extract was re-dissolved in
improving complexion, in cleaning vagina, and it is
methanol and used for the HPTLC analysis. Pre-
useful in pitta and kapha. They are used in diabetes,
coated Silica Gel F254 (Merck) plate was used for
diarrhoea, leucorrhoea, menorrhoea, nervous disorder,
stationary phase and Chloroform:Ethylacetate:Formic
and vaginal diseases. It is also widely used in the
acid (2.5:2:1.5) used as mobile phase. After
treatment of skin diseases, ulcer and soreness in the
mouth (Nadkarni, 1954; Aiyer & Kolammal, 1957; development, the plate was sprayed with 1% vanillin
Mooss, 1976; Kurup et al., 1979; Warrier, 1994; Joshi & sulphuric acid and heated at 105 °C in hot air oven for
Upadhye, 2008; Gayathri & Kannabiran, 2008). 5 to 10 min to develop the colour of the bands.
In the present study, an attempt was made to study
the comparative pharmacognostic features of the Results
barks of the above 4 species used in Nalpamaram, an Ficus racemosa L.
important Ayurvedic drug.
Macroscopic features
The bark has a thickness of about 6 to 15 mm and
Materials and methods
is grayish-green with a fairly smooth and soft surface,
Morphological and anatomical studies with minute separating papery flakes of white tissue
For the present study fresh stem barks of 4 Ficus emerging out from outer surface, no fissures and
species were collected from Cholayil Medicinal Plants homogeneously leathery texture. Inner surface light
Conservation Park (CMPCP), Velagapuram, brown (Figure 1a).
Thiruvallur district, near Chennai after Microscopic features
authentication, and voucher specimens were
preserved. Fresh barks were cut into small pieces and The transection of bark measuring about 8 mm
immediately fixed in Formalin:acetic acid:70% thickness consists of an outer periderm measuring 72
alcohol (5 mL + 5 mL + 90 mL) for 24 h and mm thick. The rest of the bark includes a secondary
dehydrated, paraffin infiltrated and embedded in wax phloem. The periderm is superficial in origin. It
using customary techniques (Johansen, 1940; Sass, consists of regular tangentially arranged thin layers of
1940). Serial transections and tangential longitudinal cells (Figure 1b). The older layers of phellem exfoliate
sections were obtained at 10-12 mm thickness with in the form of thin membrane due to separations of
rotary microtome and the sections were stained with tangential walls between successive layers of cells.
safranin and fast green. All the photomicrographs Phelloderm is evident and consists of a few layers of
were taken with a Nikon E400 microscope. cubical parenchymatous cells (Figure 1b).

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 K. BABU, S. GOKUL SHANKAR, S. RAI

PD Ficus virens Aiton

Macroscopic features
Bark is flat to curve, measuring 2 to 3 mm in
thickness. External surface ash or grayish-brown in
colour. Surface rough with numerous lenticels.
A Internal surface rough, fibrous, longitudinally striated,
Scl pale reddish, fracture, and fibrous (Figure 2a).
PR
Microscopic features
Cr TC Scl
Outer bark is represented by a narrow zone of
ST simple periderm, which is superficial in origin.
Periderm is 138 mm thick. It consists of a phellem
zone, which is 5-10 layers of cells forming thin
continuous membranes. The outermost layers of
phellem peel off as membranes of 1-cell thickness.
Phelloderm is also distinct, consisting of about 8-10
layers of cubical cells containing chloroplast or tannin.
400 μm C Along frequent places, the periderm zone invaginates
PR towards the inner side forming a sac like pouch filled
with tannin containing dead and crushed phellem
cells. Sometimes the pouch is detached from the
Phl surface and gets buried in the inner part of the bark.
In cross-sectional view this detached periderm
appears as a circular structure and this type of
200 μm D CZ 1 mm B
structure is designated as “periderm tubes” (Figures
Figure 1. Ficus racemosa: A. External features of bark, B. Cross
2b-2d).
section of bark, C and D. Portions enlarged.
Abbreviations (Figures 1-4): AP – Axial parenchyma; Cr – Crystal; Secondary phloem is clearly distinguished into
CZ – Cambial zone; Fi – Fissure; LV – Latex vessel PD – Periderm; inner broad non-collapsed phloem, which is 920 mm
Phl – Phloem; PP – Phloem protein; PR – Phloem rays; Scl –
Sclereids; ST – Sieve tube; TC – Tannin cell.
wide. This zone consists of randomly oriented sieve
tube members, tannin containing axial parenchyma
cells, and scattered groups of gelatinous fibres. The
Secondary phloem represents the broad inner bark.
outer zone has collapsed sieve tube members, dilated
It consists of a narrow zone or non-collapsed phloem,
phloem rays, and larger groups of gelatinous fibres.
which is about 360 mm broad. The non-collapsed
The dilated rays or the axial parenchyma of the
phloem has sieve tube members and axial parenchyma,
phloem remain parenchymatous, non-differentiated
which are randomly distributed. There are also small
nests of unlignified phloem fibres possessing gelatinous in sclerenchymatous elements as in other cases.
inner walls. Most of the phloem parenchyma cells contain Phloem rays are both uniseriate and multiseriate. The
tannin. Large p-protein bodies are seen in almost all sieve rays appear homocellular or heterocellular. Laticifers
tube members. The collapsed phloem zone is very wide. are not abundant both in inner and outer phloem.
The phloem rays, which start as narrow canals in the Cubical p-protein bodies are seen almost in all sieve
non-collapsed zone, dilate extensively in the collapsed tube members somewhat abutting the sieve plates.
phloem zone. The phloem parenchyma cells and phloem Prismatic calcium oxalate crystals are fairly abundant
ray cells become lignified forming sclereids. Prismatic in the axial parenchyma and ray parenchyma cells.
calcium oxalate crystals are fairly abundant in the axial They are mostly solitary in each cell. The frequency
parenchyma and ray parenchyma cells. The phloem rays of crystals increases from centre towards the
either uniseriate or multiseriate (Figures 1c and 1d). periphery.

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Comparative pharmacognostic studies on the barks of four Ficus species

Secondary phloem differentiated into inner

PD
narrow non-collapsed zone and outer broad collapsed
zone. Non-collapsed zone consists of radial files of
sieve tube members, axial parenchyma, and gelatinous
fibres. Outer collapsed phloem has dilated rays,
A
crushed obliterated sieve tube members, thick walled
and lignified fibres, and abundant tannin filled
Cr
parenchyma cells. Laticifers are fairly abundant in the
outer secondary phloem zone. Phloem rays are both
uniseriate and multiseriate. Multiseriate rays are
Cr homocellular and uniseriate rays are either
TC PR homocellular or heterocellular (Figures 3b-3f).
Ficus benghalensis L.
400 μm C Macroscopic features
Mature bark is 12-18 mm thick, grey, closely
Phl
adhered ashy white, light bluish-green or grey patches,
slightly curve, thickness varies with the age of the tree.
Surface is deeply fissured and rough due to the
presence of longitudinal and transverse row of
CZ
200 μm D 1 mm B lenticels, mostly circular and prominent, fracture
Figure 2. Ficus virens: A. External features of bark, B. Cross
short in outer 2/3 of bark while inner portion shows
section of bark, C and D. Portions enlarged. a fibrous fracture (Figure 4a).
Microscopic features
Bark differentiated into outer bark or rhytidome
Ficus religiosa L. and inner bark or secondary phloem. Outer bark
measures 288-576 mm. Width and inner bark
Macroscopic features
measures 2.9 –3.5 mm. Periderm is deeper in origin
The bark is flat or slightly curved, varying from 5 and consists of discontinuous irregular bands of
to 8 mm in thickness, outer surface is grey or ash with sequential periderm, and originates from the deeper
thin or membranous flakes and is often covered with part of the secondary phloem. Periderm consists of
crustose lichen brown or ash coloured, surface has phellem and phelloderm. Phellem cells are
shallow irregular vertical fissures and uneven due to homogeneous thin walled rectangular and suberised.
exfoliation of cork, inner surface smooth, yellowish to Phelloderm is wide and distinct. Phelloderm cells are
orange brown and fibrous (Figure 3a). turned into cubical sclereids arranged in radial files
(Figure 4b).
Microscopic features
Secondary phloem is differentiated into inner
Bark differentiated into outer thick periderm and intact non-collapsed zone, lying next to cambial zone,
inner secondary phloem. Periderm is differentiated and outer collapsed phloem zone. In the non-
into phellem and phelloderm. Phellem zone is 360 collapsed zone phloem elements occur in small
mm thick and it is wavy and uneven in transection. clusters and consist of sieve tube members,
Phellem cells are organized into thin tangential companion cells, and axial parenchyma. Tannin is
membranous layers and the older layers exfoliate in abundant in most of the axial parenchyma cells. The
the form of thin membranes. The phelloderm zone is collapsed phloem zone consists of wide dilated rays
broad and distinct. Phelloderm cells are turned into and collapsed and obliterated sieve elements (Figures
lignified sclereids. 4b and 4c).

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 K. BABU, S. GOKUL SHANKAR, S. RAI

PD

A
Scl

PR
1 mm B

ST

PR
TC

Phl

PR
1 mm C

400 μm E
Phl

ST

Cr

Phl
Cr

TC

CZ
200 μm F 1 mm D

Figure 3. Ficus religiosa: A. External features of bark, B, C and D. Cross section of bark, E
and F. Portions enlarged.

The ray cells are turned into thick walled lignified height. The sieve tube members have 288-360 mm
sclerenchyma cells in the peripheral part of the bark. height. Large vertically oblong p-protein bodies are
The phloem rays are uniseriate or multiseriate. They invariably seen abutting the sieve plates. Laticifers are
are homocellular or heterocellular. The multiseriate abundant in the inner bark. Each laticiferous canal is
rays are 72 mm in breadth and up to 900 mm in surrounded by distinct epithelial cells (Figures 4d-4f).

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Comparative pharmacognostic studies on the barks of four Ficus species

UV-254nm (Before spray) UV-366nm (Before spray)

A
Fi PR
Pd
AP

Scl

400 μm D

PR
LV Phl
A B C D A B C D
PP
Day light (after spray)
1 mm B
ST
PP

Cr AP

Scl 200 μm E

PP
Cr

LV

PR
CZ A B C D
0.5 mm C 200 μm F
Figure 4. Ficus benghalensis: A. External features of bark, B. Cross Figure 5. HPTLC profile of Ficus spp. barks: Track -A: F.
section of bark, C. Portion under polarized light, D. racemosa, Track – B: F. virens, Track – C: F. religiosa,
Tangential longitudinal section of bark, E. Portion Track – D: F. benghalensis
enlarged, F. Phloem region.

Table 1. Physico-chemical analysis of 4 Ficus spp. barks.

Class of chemical Ficus Ficus Ficus Ficus

compounds racemosa virens religiosa benghalensis

Alcohol soluble extractive 8.48 ± 0.30 2.26 ± 0.78 7.21 ± 0.92 4.43 ± 0.95

Water soluble extractive 11.27 ± 0.47 4.39 ± 0.83 15.76 ± 0.67 7.44 ± 0.86

Total ash 16.31 ± 1.45 11.97 ± 1.18 7.86 ± 0.8 5.45 ± 0.92

Acid insoluble ash 1.35 ± 0.21 2.59 ± 0.45 0.41 ± 0.03 1.21 ± 0. 23

220
 K. BABU, S. GOKUL SHANKAR, S. RAI

Table 2. Preliminary phytochemical screening of 4 Ficus spp. barks.

Class of chemical compounds Ficus racemosa Ficus virens Ficus religiosa Ficus benghalensis

Tannins + + + +
Saponins + + + +
Flavonoids + + + +
Steroids + + + +
Terpenoids + + + +
Cardiac glycosides + + + +
Alkaloids - - - -
Quinones - - - -

+ Present; - Absent

Table 3. Rf. values of HPTLC analysis of 4 Ficus spp. barks.

F.racemosa F. virens F. religiosa F. benghalensis

Under 254 nm
0.06 0.06 0.05 0.06
0.09 - - -
- - 0.14 -
- 0.2 - -
- 0.23 0.22 0.23
- 0.28 - -
- - 0.32 -
- - 0.4 -
0.48 - 0.48
- 0.64 0.65 -
0.68 - - -
- - - 0.71
0.75 - - -
0.81 0.8 - -
- - - 0.84
0.89 0.9 - 0.9
Under 366 nm
0.05 0.06 0.05 0.06
0.14 - - -
- 0.17 0.18 -
0.23 - 0.22 -
0.24 0.25 0.24 0.24
- - 0.27 -
- 0.38 0.37 -
- - 0.43 -
- 0.48 — -
- - 0.56 -
- - 0.62 0.63
- 0.76 0.76 0.76
- - - -
- - - -
0.83 0.83 0.84 0.84
0.96
Under 550 nm (After spray)
- - - 0.15
- - 0.19 -
- 0.23 0.22 0.22
0.28 0.28 0.28 0.27
- - 0.32 -
- - 0.42 -
- - 0.53 0.53
0.84 0.85 0.84 0.86
- - - 0.88

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Comparative pharmacognostic studies on the barks of four Ficus species

F. benghalensis
AU 1000
900
800 F. religiosa
700
600
500
400 F. virens
300
200
100 F. racemosa
0
0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0
Wavelength: 260 nm [RF]

F. benghalensis
AU 1000
900
800 F. religiosa
700
600
500
400 F. virens
300
200
100 F. racemosa
0
0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0
Wavelength: 366 nm [RF]

AU 1000
900
F. benghalensis
800
700
600 F. religiosa
500
400
300 F. virens
200
100
0 F. racemosa
0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0
Wavelength: 550 nm [RF]

Figure 6. HPTLC densitometer scanning profile of Ficus spp. barks.

30
25
20
% of yield

15
10
5
0
Hexane Chloroform Methanol Water
F. racemosa F. virens F. religiosa F. benghalensis

Figure 7. Successive soxhlet extractives of barks of Ficus spp. with various solvents.

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 K. BABU, S. GOKUL SHANKAR, S. RAI

Table 4. Morphological and anatomical diagnostic features of 4 Ficus spp. barks.

S.No Characters Ficus racemosa Ficus virens Ficus religiosa Ficus benghalensis

1 Thickness 8 mm 2-3 mm 5-8 mm 12-18 mm

2 Physical Soft surface with minute Hard and rough surface. Hard and rough surface often Hard and rough surface.
features papery flakes, smooth. covered with crustose lichen.

3 Colour Greyish-green colour Ash or greyish-brown. Greyish-white with Grey colour with dark patches.
with brown patches. green spots.

4 Fissure Absent. Absent. Fissures shallow, vertical Fissures deep, irregular and
and irregularly oriented. vertically oriented.

5 Lenticels Absent. Lenticels irregular Absent. Lenticels in longitudinal and

with black spot. transverse row, mostly circular
and prominent.

6 Periderm Thin measuring Thin measuring about Thick measuring about Very thick measuring about
about 72μm. 138μm and characteristic 360μm and breaking into 288-576μm and distinct.
feature is the periderm irregular flakes.
tubes.

7 Phellem Thin, membranous Thin, peel off as Thick and wavy, uneven in Thick and homogeneous thin
and easily peel off. membranes of one transaction. Older layers walled rectangular suberised
cell thickness. exfoliate in the form of thin cells.
tangential membranes.

Phytochemical studies predominance and also used in various ailments

The percentages of alcohol-soluble and water- (Sivarajan & Balachandran, 1994; Joy et al., 2001).
soluble extractives, total ash, and acid-insoluble ash Barks of some of these 4 species, such as Ficus
are tabulated in Table 1. Presence and absence of virens, are also equated with many other species like F.
different phyto-constituents were detected (Table 2). microcarpa L. f. F. infectoria Roxb., F. arnottiana Miq,
HPTLC fingerprint profiles were developed and are F. lacor Buch-Ham and F. talboti King (Nadkarni,
presented in Table 3 and Figures 5 and 6. The 1954; Singh & Chunekar, 1972; Kapoor & Mitra, 1979;
percentages of successive soxhlet extractives were Sharma, 1983). Hence, it is very difficult to identify
calculated and results are depicted in a histogram the original from the adulterants/substitutes while
(Figure 7). procuring crude drug from the market.
Discussion and conclusion Based upon the macroscopic and microscopic
Tree bark is very complex in structure and has the features and HPTLC profiles of the barks of these 4
potential of containing many primary and secondary species, we can identify them with some specific
metabolites. Products stored in the bark are useful for characters. The diagnostic features of the 4 Ficus barks
preparation of many drugs. The complex structure of are tabulated in Table 4.
the bark can be utilized for botanical identification to The barks of 4 Ficus species contains tannin, wax,
maintain the quality and purity of the drug (Brinda et saponin gluanol acetate, β-sitosterol, leucocyanidin-
al., 2000). 3 – O – β – D - glucopyrancoside, leucopelargonidin
Nālpāmaram is an important group of Ayurvedic – 3 – O – β – D - glucopyranoside, leucopelargonidin
formulation that constitutes the barks of Ksīrivrksās – 3 – O – α – L - rhamnopyranoside, lupeol, ceryl
(4 laticiferous tree species), namely Ficus racemosa, F. behenate, lupeol acetate, α-amyrin acetate,
virens, F. religiosa and F. benghalensis, widely used in leucoanthocyanidin, and leucoanthocyanin (Husain
the treatment of skin diseases with pitta and rakta et al., 1992).

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Comparative pharmacognostic studies on the barks of four Ficus species

The preliminary phytochemical screening shows studied and reported the pharmacognosy of the
that all the barks possess similar types of Ficus species individually, a relative and
phytoconstituent groups. However, significant comparative study of the species providing key
differences were observed in the physico-chemical diagnostic tools has not been done earlier. We
analysis and successive soxhlet extractions with reported in the current study that on the basis of
different solvents. several cumulative characters, the bark of the 4
Comparative HPTLC fingerprint also shows species of Ficus can be easily differentiated or
marked differences in their profiles. In UV 254 nm, distinguished from adulterants.
except 2 common bands at Rf. 0.06 & 0.48, the other
bands do not match. In UV 366 nm, all the barks
Acknowledgements
show 1 similar common band at Rf. 0.24. F. religiosa
and F. benghalensis have 2 common bands at Rf. The authors are grateful to Dr. V.P. Sidhan
0.62 & 0.76. F. virens has 1 common band at Rf. 0.76 (Chairman), Mr. V.S. Pradeep (Managing Director),
and the band at Rf. 0.62 is absent. In visible light and Mrs. Jayadevi Pradeep (Director) from Cholayil
(after spray) all the barks shows 2 similar common Private Limited for their constant support and
bands of violet colour at Rf. 0.28. F. virens, F. encouragement. The authors are also thankful to Prof.
religiosa, and F. benghalensis have 1 common band P. Jayaraman (Director), Plant Anatomy Research
of pink colour at Rf. 0.22 and this band is absent in Centre (PARC), Chennai, for the valuable guidance
F. racemosa. Though earlier researchers have during the study.

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