Impact factor: 0.3397/ICV: 4.10 248 PHARMACOGNOSTICAL ...
Transcript of Impact factor: 0.3397/ICV: 4.10 248 PHARMACOGNOSTICAL ...
Impact factor: 0.3397/ICV: 4.10 248
Krunal et al. / Pharma Science Monitor 5(3) Supl-1, Jul-Sep 2014, 248-255
Pharma Science Monitor 5(3) Supl-1, Jul-Sep 2014
PHARMACOGNOSTICAL STANDARDIZATION OF SAPTAPARNA (ALSTONIA
SCHOLARIS R. Br.) STEM BARK
Kamlesh Bhogayata1, B. R. Patel2, Jignesh Kevalia3, Bhakti Chhaya4, Bharatkumar M. Gohel5,
Krunal Doshi6* 1Assistant Professor, Dept. of Dravyaguna, Sheth J. P. Govt. Ayurved College, Bhavnagar Gujarat, India. 2Assistant Professor, Dept. of Dravyaguna, IPGT & RA, Gujarat Ayurved University, Jamnagar, Gujarat, India. 3Head, Dept. of Pharmacognosy, Indian Institute of Ayurvedic Pharmaceutical Sciences, 4Assistant Professor, Dept. of Shalakya, Sheth J. P. Govt. Ayurved College, Bhavnagar Gujarat, India. 5Assistant Professor, Dept. of Preventive and Social Medicine, PDU Govt. Medical college, Jamnagar road, Rajkot, Gujarat, India. 6Lecturer, Dept. of Dravyaguna, Indian Institute of Ayurvedic Pharmaceutical Sciences, Gujarat Ayurved University, Jamnagar.
ABSTRACT The bark of Alstonia scholaris R. Br. (Apocynaceae) locally known as ‘Saptaparna’ is reported to have anticancer, anti-helminthic, anti-diarrhoeal, anti-asthamatic, anti-malarial etc., activities. The present work represents the study carried out for quality control of herbal drugs which comprises of macroscopy, microscopy, powder characters and histochemical study of stem bark of A. Scholaris R. Br. Result showed the stem bark was dark grey to brown in colour without any odour and bitter in taste. Microscopy revealed the presence of cork, cambium, phloem, medullary rays and stone cells. Powder microscopy of the sample drug exhibited the occurrence of cork cells, stone cells, prismatic crystals, starch grains, phloem, fibers etc. The presence of starch grains, prismatic crystals, stone cells and fibers were also showed by histochemical study. The present study on pharmacognostical investigation of A. scholaris R. Br. bark will be helpful in developing standards for quality, purity and sample identification of this plant. KEYWORDS: Alstonia scholaris R. Br., Saptaparna, Stem Bark, Macroscopy, Microscopy, Histochemical study. INTRODUCTION
Pharmacognosy has become one of the pillars in areas like pharmacy, medicine, natural product
chemistry and many others allowing scientists to recognize the importance of plants as sources of
medicines. This approach has initiated active research programs either to isolate new lead
compounds or to produce standardized extracts.[1] For this it is very necessary to evaluate various
qualitative and quantitative parameters, which may be helpful in setting standards for particular
medicinal plant/parts of the plant. With the help of these standards one can easily identify and
characterize an individual drug, which may play a major role in maintaining quality and purity of
PHARMA SCIENCE MONITOR
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that particular drug and its formulation and prevent it from being adulterated by drug of same or
other genius having low potency.[2]
The present study deals with the standardization of one such medicinal plant Saptaparna-
Alstonia scholaris R. Br. is an important medicinal plant in folklore medicine. The plant belongs
to family Apocynaceae and is native to India. It grows throughout India, in deciduous and ever-
green forests and in plains. Juice of leaves and tincture of the bark acts as a powerful
galactogogue and also used in cases of snake bite. Milky juice of the plant is applied on wounds
and ulcers. The bark is bitter, acrid, astringent, digestive, laxative, thermo genic, antipyretic,
galactogogue, cardiotonic and tonic. It is useful in abdominal disorders, fevers, leprosy, skin
diseases, chronic and foul ulcers, asthma, bronchitis and helminthiasis.[3] The bark extract
induces the cellular immune response at low doses and inhibited the delayed type of
hypersensitivity reaction at high doses.[4] The methanolic extract of this plant exhibits
pronounced anti plasmodial activity.[5] The alkaloid fraction of A. scholaris was found to have
potential anticancer agent.[6] It’s bark extract showed chemo preventive potential against skin
tumor genesis in swiss albino mice.[7] Ethanolic extracts of A. scholaris possess powerful in-vitro
antioxidant activity.[8]
Barks from trees and shrubs constitute an important source of drugs used in Indian System of
Medicine. Just after collection and on drying, morphologically or organoleptically several barks
resemble each other in many respects and may exhibit few minor variations in their size, shape,
outer and inner surface, colour, mode of fracture, odour, tastes, etc. However, microscopical
characters performs the major parts and assists us greatly for the correct identity. Hence, an
attempt has been made to ensure properties of Saptaparna (Alstonia scholaris R. Br.) stem-bark
through pharmacognostical study.
MATERIALS & METHODS
Collection of Sample
Bark of Alstonia scholaris R. Br. was collected from the periphery Gujarat state by Pharmacy
department of IPGT & RA, GAU, Jamnagar. Further the authenticity of the sample was
confirmed by the experts of Gujarat Ayurved University and by comparing their characters
mentioned in various floras.
Preservation
The drug was thoroughly washed with running water and cut into small pieces and preserved in
the solution containing [Formalin: Glacial Acetic Acid: Ethanol 70 % in 5: 5: 90]. While some
pieces were dried on sun and powdered for studying its powder microscopy.
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Preparation of illustrations
The section were taken by free hand and their photo graphs were taken by using Canon digital
camera attached to Zeiss microscope and powder characters were drawn with the help of Camera
Lucida lying at the Pharmacognosy department of I.P.G.T. & R.A., Gujarat Ayurved University,
Jamnagar.
Macroscopic & organoleptic evaluation
The characters were studied systematically as per the methods described in the text books of
pharmacogosy. The specimen were observed as such with naked eyes or with the help of
dissecting microscope.[11] Various parameters of the plant material, such as size, shape, colour,
odour and taste of the stem bark were also recorded.[9, 10, 11]
Microscopic evaluation
Thin free-hand sections of the stem bark were made and washed with chloral hydrate solution.
The stain was made with phloroglucinol and conc. HCl solution. Diagnostic characters in TS and
powder of stem bark of A. scholaris R. Br. was studied with and without staining.
Microphotographs were taken using Carl Zeiss binocular microscope.[11]
Histochemical Test
Histochemical tests were performed to detect the kinds of cell wall like lignin, primary
metabolites like starch grains, ergastic substances like crystals of calcium oxalate and calcium
carbonate, etc. present in the sample.[12]
RESULT & DISCUSSIONS
Macroscopic & Organoleptic characters
Pieces are curved or flat or occasionally quilled, measuring 8 – 12cm in length, 5 - 8cm broad
and 5 – 15mm in thickness, externally uneven, transversely and longitudinally fissured, marked
with numerous patches of whitish to silver coloured lenticels; surface internally smooth,
longitudinally striated; fracture outer hard and short, inner granular; fractured surface shows
narrow cork and wide phloem zone traversed by latex canals; colour externally dark grey –
brown, internally brownish buff – dark grey, young bark is somewhat paler; odour not
characteristic; taste is bitter and slightly astringent. (Fig. 1)
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FIGURE 1 – MACROSCOPIC CHARACTERS OF INTERNAL AND EXTERNAL
SURFACE OF A. SCHOLARIS R. Br. STEM BARK
Microscopic evaluation:
Diagrammatic section: Diagrammatic TS of bark shows outer wide multi layered cork, lying
underneath the cortex and a very large region of inner phloem traversed with laticeferous canals.
Cortex: Detailed TS shows 80 – 100 rows of narrow, rectangular, thin to thick walled, yellow to
brown coloured cork cells at places thick walled with different lumen stone cells are seen, within
the cork a well-defined, thin walled, broadly rectangular, narrow cells or phellogen or cork
cambium; phelloderm or secondary cortex is thin walled, nearly cubical to rectangular, many
layered, traversed with cubical to oblong, groups of stone cells varies in size and with different
kind of lumen, sometimes with pits; phloem is very wide, parenchymatous, the cells being
discontinuous because of parallel arrangement of uni to biserriate, rectangular, thin walled
medullary rays, radially elongated in the portion nearer the wood but becomes broad towards the
distal ends, laticiferous canals traversed throughout the phloem region; prismatic crystals of
calcium oxalate and simple, spherical starch grains scattered throughout the section. (Fig. 2)
Partly cut TS of the bark of Alstonia scholaris R. Br. shows cork, cortex, and prismatic crystals along with starch grains. (Blue colour shows presence of starch grains).
Partly cut TS of the bark of Alstonia scholaris R. Br. shows cortex region with the arrangement of stone cells.
Partly cut TS of the bark of Alstonia scholaris R. Br. showing lignification of stone cells in cortex
region.
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FIGURE 2 – MICROSCOPIC CHARACTERS OF A. SCHOLARIS R. Br. STEM BARK
Powder microscopy
A light brown powder with fine texture and without any characteristic
odour having bitter taste has following distinguished microscopical
characters:
1. The abundant groups of stone cells of various sizes, shapes and thickness
with distinct radiating pits and striations. 2. Plenty of sclereids with highly
thickened and striated walls with various sizes and shapes. 3. The abundant prismatic crystals of
calcium oxalate scattered as such and embedded in parenchymatous cells of phelloderm, phloem
and stone cells. 4. The fragments of parenchymatous tissue and tangentially- longitudinally cut
medullary rays of phloem, embedded with lactiferous canals which at places show bulging
enlargements and are filled with granular contents. 5. The fragments of cork in surface view and
in sectional view; in surface view the cells are thick walled, hexagonal to pentagonal in shape
while in sectional view, the cells are thick walled, rectangular to squarish in shape, often with
lignified and pitted walls. 6. The fragments of beaded hexagonal to elongated, lignified
parenchymatous cells of the phloem. 7. Simple, spherical to oval, starch grains scattered as such
and embedded in the parenchymatous cells. 8. Thick walled, striated, non lignified fibres
occasionally present tin the young bark only. (Fig. 3)
Partly cut TS of the bark of Alstonia scholaris R. Br. showing lignification
of stone cells in cortex region (enlarge view)
Partly cut TS of the bark of Alstonia scholaris R. Br. shows cortex region with broad parenchyma containing abundant starch grains and groups of 5-7 rows of
stone cells.
Partly cut TS of the bark of Alstonia scholaris R. Br. shows broad zone of
phloem consisting phloem parenchyma, phloem fibres, medullary
rays and latex cells.
Longitudinally and tangentially cut arrangement of medullary rays in
TLS view.
Phloem region in large view shows phloem parenchyma traversed with prismatic
crystals of calcium oxalate and starch grains.
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FIGURE 3 – POWDER CHARACTERSTICS OF A. SCHOLARIS R. Br. STEM BARK
TABLE NO. 1 - COMPARATIVE STATEMENT OF STEM BARK AND ITS POWDER
FORM
Sr. Characters Stem bark of
A. scholaris R. Br.
Powder of
A. scholaris R. Br.
1. Taste Astringent & Bitter Bitter
2. Color Grayish brown (Ext.)
Light brown Creamiest white (Int.)
3. Odor Not characteristic Not characteristic
4. Surface Rough & Uneven (Ext.
Texture – fine Smooth (Int.)
Histochemical characters
1. Test for Starch grains:
Mount a section of sample in water, add a drop of Iodine, blue colour shows the presence of
starch grains.
2. Test for Crystals of Calcium oxalate:
Cork in sectional view Cork in surface view Part of phloem in
tangential - section
Prismatic crystals, Starch grains,
Parenchyma filled with prismatic crystals.
Beaded phloem parenchyma,
Longitudinal cut fragments of laticiferous
canal
Fibers Sclereides
Stone cells
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They are insoluble in glycerin and phenolic reagents but soluble in most acids commonly used
hydrochloric acid. By adding2 - 3 drops of hydrochloric acid crystals of calcium oxalate gets
dissolved.
3. Test for fibres and stone cells:
Fibres of Sclerenchyma and stone cells gives pink to red colour when stain with Phloroglucinol
and HCl.
CONCLUSION
Standardization of herbal drugs is a topic of great concern because of the great variability
derived from heterogeneous sources. Alstonia scholaris R. Br. is a plant with large medicinal
advantages and it’s stem bark plays a major role in these medicinal properties. Thus the present
study will provide referential information for correct identification and standardization of this
plant material through pharmacognostical study as basic tool for identification.
REFERENCES
1. Ameenah and Gurib-Fakim: Review medicinal plants traditions of yesterday and drugs of
tomorrow. Molecular Aspects of Medicine. 2006; 27:1–93.
2. Doshi KA, Acharya RN, Harisha CR and Preeti P: Pharmacognostical evaluation of the
wild and cultivated variety of Eranda (Ricinus communis Linn.) root. Phcog J 2012; 4:
54-58.
3. Kirtikar KR and Basu BD: Indian Medicinal Plants. Lalit Mohan Basu Publishers,
Allahabad, India, Vol. I, 2002: 111-114.
4. Iwo MI, Soemardji AA, Retnoningrum DS and Sukrasno UUM: Immunostimulating
effect of pule (Alstonia scholaris L. R. Br., Apocynaceae) bark extracts. Clinical
emorheology and Microcirculation 2000; 23: 177-183.
5. Keawpradub N, Kirby GC, Steele JCP and Houghton PJ: Antiplasmodial activity of
extracts and alkaloids of three Alstonia species from Thailand. Planta Medica 1999; 65:
690 - 694.
6. Jagetia GC and Baliga MS: Evaluation of anticancer activity of the alkaloid fraction of
Alstonia scholaris (Sapthaparna) in vitro and in vivo. Phytotherapy Research 2006; 20
(2): 103-109.
7. Baliga MS: Alstonia scholaris Linn R Br in the Treatment and Prevention of Cancer:
Past, Present, and Future. Integrative Cancer Therapies 2010; 9 (3): 261-269.
Impact factor: 0.3397/ICV: 4.10 255
Krunal et al. / Pharma Science Monitor 5(3) Supl-1, Jul-Sep 2014, 248-255
8. Arulmozhi S, Mazumder PM, Ashok P and Narayanan LS: In vitro antioxidant and free
radical scavenging activity of Alstonia scholaris Linn. R. Br. Iranian Journal of
Pharmacology and Therapeutics 2007; 6: 191-196.
9. Kokate CK, Purohit AP and Gokhale SB. Pharmacognosy. Nirali Prakashan, Pune,
Edition 2, 2008: 63.
10. Anonymous: Ayurvedic Pharmacopoeia of India. Govt. of India, Ministry of Health of
Family Welfare, New Delhi, Edition 1, Vol-2, Part-2, 2008: 15-17.
11. Khandelwal KR: Practical Pharmacognosy - Techniques and Experiments. Nirali
Prakashan, Pune, Edition 9, 2002: 24-29,149-156.
12. Krishnamurthy KV: Methods in the Plant histochemistry. Vishwanandhan Pvt Limited,
Madras, 1988: 1-77.
For Correspondence Krunal Doshi Email: [email protected]