13 | P a g e International Standard Serial Number (ISSN): 2319-8141

International Journal of Universal Pharmacy and Bio Sciences 5(2): March-April 2016

INTERNATIONAL JOURNAL OF UNIVERSAL

PHARMACY AND BIO SCIENCES
IMPACT FACTOR 2.093***
ICV 5.13***
BIO SCIENCES RESEARCH ARTICLE …………!!!

AN ANTI INFLAMMATORY FOLK LORE HERB

BULBOPHYLLUM NEILGHERRENSE WIGHT.- A SOURCE OF
CALCIUM
VRINDA G NAIR1*, K. NISHTESWAR2, VIDHYA UNNIKRISHNAN3, PREETI
PANDYA4
1
PG Scholar, Dept. of Dravyaguna, I.P.G.T&.R.A, Jamnagar.
2
Ex.Professor & HOD, Dept. of Dravyaguna, I.P.G.T&.R.A, Jamnagar.
3
Ph.D Scholar, Dept. of Dravyaguna, I.P.G.T&.R.A, Jamnagar.
4
Pharmacognosy, Dept. of Dravyaguna, I.P.G.T&.R.A, Jamnagar.

KEYWORDS: ABSTRACT

Bulbophyllum Bulbophyllum neilgherrense Wight is an epiphytic orchid found in the

neilgherrense, Western Ghats with a few important ethno medicinal claims.

Pseudobulb, Calcium
Pharmacognostical study of its pseudobulb and leaf along with calcium
estimation, heavy metal
analysis. estimation were carried out. Microscopic evaluation of leaf showed the

For Correspondence: presence of mesophyll tissue filled with chloroplast pigments, which is
VRINDA G NAIR *
spirally thickened. Microscopy of pseudo bulb showed thick waxy cuticle
Address:
PG Scholar, Dept. of followed by epidermis, oil globules, starch grains, tracheids, fibers etc.

Dravyaguna, Both of them showed presence of plenty of acicular, prismatic and rosette
I.P.G.T&.R.A, Jamnagar.
crystals of calcium oxalate, raphides, mucilage cells although more were

found in leaves. This paper discusses the macroscopical, microscopical

features, calcium estimation and heavy metal analysis of pseudobulb and

leaf of the plant.

Full Text Available On www.ijupbs.com

 14 | P a g e International Standard Serial Number (ISSN): 2319-8141

INTRODUCTION:
Ethno medicine has been widely appreciated for its treasure of knowledge. Research on various ethno
medicinal practices is on demand since long for the discovery of new chemical compounds or
medicines. They give new hope for treatment of some diseases which are incurable by other systems
of medicine. Bulbophyllym neilgherrense Wight. is one of those ethno medicinal plants which is
widely used by the tribes of Western Ghats for prevention as well as treatment of many diseases. 1 It is
an epiphyte belonging to the family Orchidaceae. The special characteristic of the plant is the
presence of green, angled pseudobulb, which is a solid bulbous enlargement of the stem to preserve
water and nutrients. Pseudobulbs in this orchid are 4cm long and 2cm across, smooth, green and four
angled. Leaves 10cm-15cm long, 2-3cm broad, coriaceous, elliptic to broadly oblong, obtuse at apex,
base narrowed tapering into short petiole attached to the pseudobulb. 2,3 Its pseudobulb, leaf and whole
plant are used by traditional practitioners. Pseudobulb is used by the Gowlis of Uttara Kannada
district of Karnataka to cure different ailments like heart diseases, rheumatism, leucoderma and as a
tonic.4-6 Paste of pseudobulbs is externally applied for arthritis and used as a secondary ingredient
with Gymnema sylvestre R. Br. and other herbal ingredients in Type 2diabetes with ailments of back
pain and sore on sacral region. 7
Materials and Methods:
Whole plant was collected from its natural habitat in Vellingiri Hills,Coimbatore,Tamil Nadu, India
during January 2016. Botanical identity was confirmed at Pharmacognosy Laboratory, IPGT&RA,
Gujarat Ayurveda University, Jamnagar. A herbarium specimen is preserved in the pharmacognosy
laboratory IPGT&RA, Gujarat Ayurved University, Jamnagar (voucher specimen number 6154/2016)
Pseudobulbs were collected, cut into small pieces and dried in shade. Dried material was ground and
sieved in mesh 40-60#. Macroscopic evaluation of fresh pseudobulb was done. Size, shape, colour and
texture were noted in detail. Organoleptic study of the pseudobulb powder was carried out for
determining its colour, odour, taste and texture. 8
For microscopic evaluation, free hand sections of the fresh sample were taken and washed with
chloral hydrate solution. Sections were first observed in distilled water, later stained with
phloroglucinol and conc. HCl. Pseudobulb powder was viewed under microscope for the detection of
various cells and their contents. Photographs were taken by Carl zeisstrinocular microscope. 9,10 Thick
sections were treated with various reagents to locate chemical constituents such as tannin, mucilage,
crystals and tissue structures like lignin etc. in histochemical tests. 11
Preliminary physicochemical and Phytochemical analysis of pseudobulb and leaf were carried out
following standard procedure of API.

Full Text Available On www.ijupbs.com

 15 | P a g e International Standard Serial Number (ISSN): 2319-8141

Calcium estimation and heavy metal analysis were carried out in SICART, Sardar Patel Centre for
Science and technology, Anand, Gujarat.
Results and Discussion
Pseudobulb
Macroscopic characters (Fig.1)
Pseudobulbs 3cm-5cm long and 2cm across, smooth, green, globular-conical when young, conical-
ovoid and angled when matured, 4 angled, bearing a single leaf at its apex. Angles gradually may
expand to wings as seen when dried. Pseudobulbs get yellowing with age. They remain 3-5cm apart
on the creeping rhizome. Pseudobulbs are rich in mucilage.
Organoleptic characters:-Color –green when young and greenish yellow when matured, odour-Not
Characteristic, Taste- Sweet, slimy, Touch – smooth.

Fig.1 Macroscopic characters

Microscopical characters:
Transversely cut section of the pseudobulb is tetrangulared to circular in outline, with cuticularised
epidermis and number of vascular bundles distributed all over the ground tissue. [Fig.2A] Detailed
section shows the outer epidermis made up of single layered oval, compactly arranged parenchyma
cells interrupted by the presence of sunken stomata at some places. Epidermis is covered with thick
waxy cuticle. Hypodermis is 3-6 layered with radially irregular walls of simple parenchyma
cells.[Fig.2B] Cortical parenchyma appears like honey comb structure with thin walled parenchyma
followed with a thick band of sclerenchma fibres surrounding the ground tissue. Ground tissue is
mostly made of loosely arranged parenchyma cells with scatteredly arranged vascular bundles. The
vascular bundles are smaller in size and more in number towards the periphery and lesser in number
with bigger size towards the centre. Each vascular bundle is conjoint collateral and closed. [Fig.2C]
Parenchyma cells are pitted, lignified or beaded with mesh like network. Mucilage containing cells,

Full Text Available On www.ijupbs.com

 16 | P a g e International Standard Serial Number (ISSN): 2319-8141

starch grains, raphides of acicular crystals and yellowish brown content found throughout the section
mostly while prismatic crystals of calcium oxalate found rarely.

Fig.2A Fig.2B Fig. 2 C

Powder analysis:
Organoleptic evaluation of the sample powder showed brownish colour, with no characteristic smell.
On touch, the powder is fibrous and on putting into mouth it is mucilaginous without characteristic
taste.
Diagnostic characters detected under microscope were annular, spiral and scalariform thickened
vessels, tracheids and fibres [Fig.3A], acicular crystals [ Fig.3B ], fragments of pitted vessel, lignified
fibres with wide lumen [Fig.3C ], mucilage cells, lignified parenchyma cells with oil
globules,[Fig.3D] simple starch grains [Fig.3E ], pitted parenchyma cells , epidermal cells in surface
view [Fig. 3F ], tannin content material.

Fig. 3A Fig. 3B Fig.3 C

Fig. 3 D Fig.3E Fig. 3F

Full Text Available On www.ijupbs.com

 17 | P a g e International Standard Serial Number (ISSN): 2319-8141

Leaf
Macroscopical characters:
Description of leaf- Leaves are 8cm-15cm long, 2-3cm broad, coriaceous, elliptic to broadly
oblong, flattened, succulent, obtuse at apex with narrow base. Midrib is prominent in the ventral
surface where as grooved on the dorsal surface. Leaf margins are simple. Leaf blade has parallel
venation, tapers into short petiole attached to the pseudobulb. A single leaf emerges from the top of
each pseudobulb.
Microscopical characters:
Leaf is dorsiventral in nature.[Fig.4A] Mesophyll tissue is filled with chloroplast pigments
consisting number of secretary cells. Vascular bundles are centrally located. Upper epidermis is
single layered with compactly arranged barrel shaped cells. These epidermal cells are covered with
thick, ridged cuticle. Lower epidermis is thickly cuticularized. Both the epidermis possesses sunken
stomata. Mesophyll is spirally thickened near the epidermis with banded parenchyma cells. Rest of
the mesophyll tissue is filled with undifferentiated parenchyma cells containing numerous
choloroplast, and is compactly arranged. The mesophyll also consists of some mucilage cavities.
[Fig.4B]The mesophyll also consists of raphide idioblast. In midrib portion there is a centrally
located large vascular bundle and smaller vascular bundles are passing through the main nerves.
Vascular bundle consists of phloem towards lower epidermis, xylem towards upper epidermis with
few xylem elements with xylem parenchyma and its fibres, where as phloem with few sieve
elements and fibres[Fig.4C]. Some of the mesophyll parenchyma consists of reddish brown colour
contents. The thick cuticle and sunken stomata shows the xerophytic nature of the plant, where as
the parenchyma cells, large mucilage cells show the hydrophytic nature of the plant.

Fig.4A Fig.4B Fig.4C

Powder analysis:
Diagnostic characters of powder shows stomata from epidermis, fibres,[Fig.5A] plenty of acicular,
prismatic and rosette crystals of calcium oxalate, [Fig.5B], raphides, mucilage cells, spiral vessels,

Full Text Available On www.ijupbs.com

 18 | P a g e International Standard Serial Number (ISSN): 2319-8141

annular thickened parenchyma cells, tannin from mesophyll tissue and epidermal cells in surface
view.[Fig.5D]

Fig.5A Fig.5B Fig.5C

ANALYTICAL STUDY:

1] PHYSICO-CHEMICAL ANALYSIS

S.No Parameters Pseudobulb Leaf

1. Loss on drying % w/w 4.045 3.844
2. Total Ash Content% w/w 4.2 6.6
3. Acid Insoluble Ash % 0.01 0.09
w/w
4. Water Soluble Extractive 33 36
Value % w/w
5. Alcohol Soluble 10 12
Extractive Value % w/w
6. pH 5.5 5.5

Above table shows the results of various physico-chemical tests conducted on officinal part
i.e., pseudobulb and leaf of B.neilgherrense plant.

Full Text Available On www.ijupbs.com

 19 | P a g e International Standard Serial Number (ISSN): 2319-8141

2) QUALITATIVE ANALYSIS

S.No Parameters Test performed Pseudobulb Leaf

1. Alkaloids (M.E.) Dragendorff‘s + +
reagent
2. Alkaloids (W.E.) Dragendorff‘s + -
reagent
3. Carbohydrates (W.E.) Molisch test + -
4. Saponin Glycosides Foam test + +
(W.E.)
5. Tannins & Phenols Ferric chloride + +
(M.E.) solution test
6. Tannins & Phenols Ferric chloride + +
(W.E.) solution test
7. Flavanoid (M.E.) Shinoda's test + +
8. Steroids (M.E.) Sarkowski reaction + +
9. Reducing sugar(W.E.) Fehling's test + +
10. Amino acids (W.E.& Biuret test - -
M.E.)

ICP test report

S.No Sample ID Test Testing Name of the Results
parameter method/procedure instrument

1. Whole plant Lead, ppm In house ICP-OES 1.7345

powder
Mercury,ppm BDL*

Arsenic, ppm 0.5680

Calcium,ppm 6697.1

2. Leaf powder Calcium, In House ICP-OES 10042.0

ppm

3. Pseudo bulb Calcium, In House ICP-OES 7685.3

powder ppm

*Below detection level

Microscopic evaluation of leaf and pseudobulb showed the presence of many acicular, prismatic
and rosette crystals in leaf as well as pseudobulb and based on this finding calcium estimation was
carried out. The calcium values were found to be 6697.1ppm in whole plant, 10042.0 ppm in leaf
and 7685.3 ppm in pseudobulb. Leaves of B.neilgherrense Wight. showed qualitatively and

Full Text Available On www.ijupbs.com

 20 | P a g e International Standard Serial Number (ISSN): 2319-8141

quantitatively more calcium content (10042.0ppm) in comparision to pseudobulb (7685.3ppm).

Heavy metal like lead, mercury and arsenic were found within the permissible limits.
Preliminary physico-chemical and qualitative analysis showed water soluble and alcohol soluble
extractives more in leaves compared to pseudobulb. Because of the mucilaginous nature of pseudo
bulb loss on drying was found to be higher in pseudobulb. pH was same in two samples. Ash value
was found to be more in leaves which may be due to the presence of more inorganic matter
(calcium oxalate crystals).
Water extract of pseudobulb and leaf showed the presence of saponin glycosides, tannin ,phenols
and reducing sugar. Methanol extracts of both showed alkaloids, tannins, phenols, flavanoids, and
steroids. Carbohydrate was present only in pseudobulb water extract.
Calcium is an essential mineral required for bone formation and its proper function. It is an
important component of a healthy diet and a mineral necessary for life. The National Osteoporosis
Foundation says, "Calcium plays an important role in building stronger, denser bones early in life
and keeping bones strong and healthy later in life, other important uses being some exocytosis,
especially neurotransmitter release, and muscle contraction.
Various experimental studies were carried out to validate role of calcium in anti inflammatory
activity. One among them provided ample evidence that co-administration of Calcium gluconate
along with anti-inflammatory drug Etoricoxib, enhanced its anti inflammatory action in acute as
well as subacute models of inflammation. Earlier calcium chloride was advocated for the treatment
of urticaria, acute edema, pruritus and erythema. Calcium carbonate & calcium gluconate were used
for the treatment of insect stings and calcium hydroxide to suppress periapical inflammation in
dental practice.12 In another experimental study which was carried out with calcium carbonate and
calcium gluconate in two groups i.e with and without aspirin. Both calcium carbonate and calcium
gluconate could not show significant anti inflammatory activity on their own. Aspirin at sub
inflammatory dose when co-administered with these calcium salts showed significant anti
inflammatory response which was comparable with anti inflammatory response of aspirin at
therapeutic dose.13
CONCLUSION
Bulbophyllum neilgherrense Wight is an epiphytic orchid endemic to Western Ghats. Leaves of the
plant showed qualitatively and quantitatively more calcium content (10042.0ppm) in comparision
to pseudobulb (7685.3ppm). The above findings indicate that the plant is a good source of calcium.
Some previous studies show that calcium has significant role in anti inflammatory activity. This

Full Text Available On www.ijupbs.com

 21 | P a g e International Standard Serial Number (ISSN): 2319-8141

information suggests that the plant may be given as a calcium supplement in conditions like
osteoarthritis associated with osteoporosis.
REFERENCES:-
1. Rajendran A, Rama Rao N, Ravikumar K and Henry A N. Some medicinal orchids of
Southern India. Ancient Science of Life 1997;17 (1).
2. Abraham and Vatsala. Introduction to Orchids. Trivandrum:Tropical Botanic Garden
and Research Institute;1981.
3. Bhat Gopalakrishna K. Flora of Udupi. Udupi: Indian naturalist; 2003.
4. J.K.Maheswari ,Ethnobotany in South Asia J.Econ.Taxon. Bot. Additional series 12
scientific publishers. Jodhpur(India) (1996) : 244-249
5. Rajendran A,Rama Rao N, Ravikumar K, and Henry A N ,Some medicinal orchids of
Southern India, Ancient science of Life, (1997) ; 17(1):10–14
6. Hossain Mohammad Musharof ; Therapeutic Orchids. Traditional uses and recent
advances an overview. Fitoterapia. (2011) ; 82(104): 104
7. S.K.Kapur and Paviter Singh, RRL; Traditionally important medicinal plants of
Udhampur district Jammu Province Part I; J.Econ.Taxon.Bot.Additional series12,
Scientific Publishers, Jodhpur,1996; pg:75-81[80]
8. Kokate C K, Purohit AP, Gokhale SB. Pharmacognosy (42nd Edn). Pune: Nirali
Prakashan; 2008.
9. Trease G. E and Evans W.C. Pharmacognosy (16th Edn). Saunders: Elsevier; 2009.
10. Wallis T.E. Text book of Pharmacognosy (5th Edn). New Delhi: CBS Publishers;
1985.
11. Krishnamurty K.V; Methods in the plant histochemistry. Madras: Vishwanadhan Pvt
Limited; 1988.
12. Sonali ; An Experimental Study to Assess the Effect of Addition of Calcium on the
Anti-Inflammatory Activity of Etoricoxib; Journal of Pharma Research; ISSN: 2319-
5622.
13. Choksi Krishna et al., Calcium enhances anti inflammatory activity of Aspirin;
International Research Journal of Pharmacy ; IRJP 2(3)2011; 82-85; ISSN 2230-
8407.

Full Text Available On www.ijupbs.com