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Phytochemical, proximate and mineral compositions of Bryophyllum

Pinnatum (Never die) medicinal plant Odangowei I Ogidi, Ngozi G Esie and
Oluchi G Dike

Article · January 2019

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 Journal of Pharmacognosy and Phytochemistry 2019; 8(1): 629-635

E-ISSN: 2278-4136
P-ISSN: 2349-8234
JPP 2019; 8(1): 629-635 Phytochemical, proximate and mineral
Received: 07-11-2018
Accepted: 08-12-2018 compositions of Bryophyllum Pinnatum (Never
Odangowei I Ogidi
die) medicinal plant
Department of Biochemistry,
Federal Polytechnic, Ekowe,
Bayelsa State, Nigeria Odangowei I Ogidi, Ngozi G Esie and Oluchi G Dike
Ngozi G Esie Abstract
Department of Biotechnology,
Bryophyllum pinnatum is an indigenous and exotic plant used widely by the traditional practitioners for
Federal University of
treating various ailments like renal calculi, hypertension, asthma, cold, abscesses, bleeding disorders.
Technology, Owerri, Imo State,
Nigeria
Phytochemical, proximate and mineral compositions of leaves, stems and roots of Bryophyllum pinnatum
(Never die) plant was analyzed using the standard method of Association of Analytical Chemist (AOAC)
Oluchi G Dike and Atomic Absorption Spectrophotometeric (AAS) method. The qualitative phytochemical result
Department of Science showed that plant samples contains alkaloids, tannins, saponin, terpenoid, glycoside, phenols and
Laboratory Technology, Federal flavonoid which was absent in the roots of the plant. Quantitative phytochemical results ranges from
Polytechnic, Ekowe, Bayelsa alkaloid 18.72 ± 5.70%, tannins 12.40 ± 3.64%, saponin 2.46 ± 1.20%, flavonoid 2.36 ± 0.98%,
State, Nigeria terpenoid 3.77 ± 2.88% and phenol 10.48 ± 8.59%. Proximate results ranges from (3.18± 3.13%)
moisture, (1.87 ± 1.81%) ash, (3.79 ± 2.96%) protein, (0.73 ± 0.56%) lipid, (3.10 ± 1.84%) fiber, (96.87
± 96.81%) dry matter and (92.35 ± 90.90%) nitrogen free element (NFE). Mineral results ranges from
40.88 ± 28.65ppm, 36.56 ± 18.53ppm, 48.72 ± 29.78ppm, 30.4 ± 17.17ppm, 2.339 ± 1.489ppm, 0.27 ±
0.20ppm, 0.20 ± 0.12ppm, 0.087 ± 0.033ppm, 0.66 ± 0.40ppm for Ca, Mg, Na, K, Fe, Mn, Cu, Zn and
PO4 respectively. The presence of these phytochemicals, proximate and mineral elements in the plant
could be part of the contributing factors which suggest the use of the plant for various therapeutic
applications. This also indicates that Bryophyllum pinnatum plant is a good source of human nutrition
and should be included as dietary supplement.

Keywords: Bryophyllum pinnatum, phytochemicals, proximate, minerals, medicinal plant

Introduction
Plants play important roles in discovery associated with new beneficial therapeutic agents and
have received significant focus because of their bio- active substances like antioxidants,
hypoglycemic and hypolipidemic factors. Plants have invariably been exemplary source of
drugs and a number of currently available drugs happen to be derived directly or indirectly
from them. This natural source has received considerable attention for discovery and
development of leads as new drug molecules, because of its diversity. Rural people depend on
herbal and traditional medicines to cure their diseases as medicinal plants are easily available
in their surroundings and have low cost with increased efficacy and reliability [1]. According to
World Health Organization (WHO) 80% of the population rely on traditional medicine as a
source of primary health care needs [2].
The use of traditional medicines and medicinal plants in mainly developing countries as
remedial agents for the maintenance of health has been broadly observed [3]. Modern-day
pharmacopoeia however contains at least 25% drugs derived from plants and many others,
which are synthetic analogues, built on prototype chemical substances isolated form plants.
Involvement in medicinal plants as a re-budding health assistance has been fuelled with the
rising charges of prescription drugs in the safeguarding of personalized health and well being
and the bio prospecting of new plant derived drugs [4]. Medicinal herbs are a source of
chemical compounds such as alkaloids, glycosides, saponin, oleoresins, sesqueterpine,
lactones and oils [5]. These biologically active ingredients are used for the prophylactic
purposes and for the different infectious diseases [6]. Due to the presence of medicative
properties, medicinal plants have been used in wide area of the world. Many diseases like
malaria, epilepsy, diarrhea, dysentery, fungal and bacterial infections have been treated by
folklore medicines [7].
Correspondence Bryophyllum pinnatum belongs to the family Grassulaceae an erect, succulent, perennial shrub
Odangowei I Ogidi that grows about 1.5m height and reproduced from seeds and also vegetatively from leaf
Department of Biochemistry, bubbils [8]. It is an introduced ornamental plant that is now growing as weed around plantation
Federal Polytechnic, Ekowe,
Bayelsa State, Nigeria
crops [9]. Bryophyllum pinnatum is commonly known as air plant, never die, miracle leaf, love
~ 629 ~
Journal of Pharmacognosy and Phytochemistry

plant. It is used in folk medicine in tropical Africa, tropical Test for alkaloids
America, India, China, and Australia. It is well known for its Extract was dissolved in dilute HCl and filtered. Filtrates were
wound healing and haemostatic properties. Traditionally, it is treated with Mayer`s reagent (potassium mercuric iodide).
used for medicinal purpose for treatment of various ailments Formation of a yellow coloured precipitate indicates the
viz. anthelmintic, immunosuppressive, hepatoprotective, anti- presence of alkaloids.
nociceptive, anti-inflammatory, anti-diabetic,
nephroprotective, antioxidant, antimicrobial, analgesic, Quantitative Phytochemical Analysis
anticonvulsant, neuropharmacological and antipyretic Depending on the above qualitative results the quantitative
activities [10]. In South Eastern Nigeria, this herb is used to assay is carried out for Alkaloids, Tannins, Phenols, Saponin,
facilitate the dropping of the placenta of a newly born baby [9]. Flavonoids and Terpenoids
The plant leaf is mildly exposed to heat and the juice
extracted and applied to the baby’s placenta on daily basis. Total Tannins Content Determination
The crushed leaves as well as the extracted juice are mixed The tannins were determined by slightly modified Folin and
with palm oil and rubbed on abscesses [8]. It is usually applied Ciocalteu method. Briefly, 0.5 ml of sample extract is added
externally. with 3.75 ml of distilled water and added 0.25 ml of Folin
The aim of this study is to determine the phytochemical, Phenol reagent, 0.5 ml of 35% sodium carbonate solution.
proximate and mineral compositions of Bryophllum pinnatum The absorbance was measured at 725 nm. Tannic acid
medicinal plant. dilutions (0 to 0.5mg/ml) were used as standard solutions. The
results of tannins are expressed in terms of tannic acid in
Materials and Methods mg/ml of extract.
Collection of Plant material
Bryophyllum pinnatum plant was collected from vegetative Total Phenol Content Determination
garden in Yenagoa, Bayelsa State. The plant parts were The phenols were determined by slightly modified Folin and
sundried, pulverized and stored in an airtight container for Ciocalteu method. Briefly, to the 200μl of the sample extract,
laboratory analysis. 800 μl of Folin Ciocalteu reagent mixture and 2 ml of 7.5%
sodium carbonate added. The total content is diluted to 7
Qualitative Phytochemical Screening volumes with distilled water and finally kept the tubes for 2
Phytochemical screening of the extracts was carried out by a hrs incubation in dark. The absorbance was measured at 765
procedure that was based on those earlier reports by [11-12, 7]. nm. Gallic acid dilutions were used as standard solutions. The
results of phenols are expressed in terms of Gallic acid in
Test for saponins mg/ml of extract.
To 0.5g of extract, 5ml of distilled water was added in a test
tube and the solution shaken vigorously and observed for a Total Alkaloid Content Determination
stable persistent froth. The frothing was mixed with 3 drops 40 ml of 10% acetic acid in ethanol was added to 1g of
of olive oil and shaken vigorously after which it was observed powdered sample, covered and allowed to stand for 4 hours.
for the formation of an emulsion. The filtrate was then concentrated on a water bath to get 1/4th
of its original volume. Concentrated ammonium hydroxide
Test for terpenoids was added drop wise to the extract until the precipitation was
0.5g of the extract was dissolved in 1ml of chloroform and complete. The whole solution was allowed to settle and
1ml acetic anhydride added, followed by the addition of 2ml collected precipitate was washed with dilute ammonium
of concentrated H2SO4. Terpenoids was indicated by hydroxide and then filtered. The residue was dried and
formation of reddish violet colour. weighed.

Test for tannins Total Flavonoid Content Determination

About 0.5g of the extract was boiled in 10ml of water in a test The total flavonoids content of samples was determined by
tube and then filtered. A few drops of 0.1% ferric chloride following the Aluminum chloride method. Plant concentrate
was added and the solution observed for brownish green or a was mixed with distilled H2O and NaNO2 solution. After 6
blue-black colouration. min, AlCl3 solution was added and enabled to stand for 6 min,
NaOH solution was added to the mixture. Immediately
Test for cardiac glycosides (keller-killiani test) distilled H2O was added to bring to the final volume and then
To 0.5g of extract dissolved in 5ml water was added 2ml of the mixture was extensively mixed and enabled to stand for
glacial acetic acid solution containing one drop of ferric another 15 min. Optical density of the mixture was recorded
chloride solution. This was underlayed with 1ml of at 510 nm. Rutin was used as a standard compound for the
concentrated H2SO4. A brown ring at the interface indicated evaluation of total flavonoid. The total flavonoids were
the presence of deoxysugar characteristics of cardenolides. A calculated using the standard curve, and expressed as rutin
violet ring may appear below the brown ring while in the equivalent in mg/g of extracts.
acetic acid layer a greenish ring may form just above the
brown ring and gradually spread throughout this layer. Total Saponin Content Determination
Test extract were dissolved in 80% methanol, 2ml of Vanilin
Test for flavonoids in ethanol was added, mixed well and the 2ml of 72%
Dilute ammonia (5ml) was added to a portion of an aqueous sulphuric acid solution was added, mixed well and heated on
filtrate of the extract. Then, concentrated sulphuric acid (1ml) a water bath at 600c for 10min, absorbance was measured at
was added. A yellow colouration indicated the presence of 544nm against reagent blank. Diosgenin is used as a standard
flavonoids. material and compared the assay with Diosgenin equivalents.

~ 630 ~
Journal of Pharmacognosy and Phytochemistry

Total Terpenoid Content Determination Estimation of crude Lipid (Ether extract)

The extract (1g) was marcarated with 50 ml of ethanol and Five gm of dry sample was weighed on a piece of glazed
filtered. To the filtrate (2.5 ml), 2.5 ml of 5% aqueous paper and transferred into an extraction thimble. The thimble
phosphomolybdic acid solution was added and 2.5 ml of was introduced into soxhlet extractor over a pad of cotton
concentrated H2SO4 was gradually added and mixed. The wool, so that top of the thimble is well above the top of the
mixture was left to stand for 30 min and then made up to 12.5 siphon. A clean dry flask was taken, weighed and was fitted
ml with ethanol. The absorbance was taken at 700 nm. with the extractor. Ether was poured along the side of the
extractor until it begins to siphon off. Then another half-a
Methods for Proximate Analysis siphonful of ether was added. The equipment thus assembled
The dry matter, moisture, ash, crude fat, crude protein with the flask was placed on a water bath at 60-80°C and the
(nitrogen x 6.25) and crude fibre contents were determined in extractor was connected with the condenser. Cool water
powdered Bryophyllum pinnatum and Vernonia amygdalina circulation was started in the condenser and allowed the
plants using the standard methods of the Association of extraction for 8 hr. Then the thimble with the material was
Official Analytical Chemists [13] while Dry Matter and removed from the extractor. The apparatus was assembled
Nitrogen Free Element contents was calculated based on the again and heated on a water bath to recover all the ether from
net difference between the other nutrients and the total the receiver flask. The receiver flask was disconnected and
percentage composition. dried it in a hot air oven at 100°C for 1 hr, cooled and
weighed.
Estimation of ash
About 2g of the sample was weighed and taken in a vitreosil (Wt. of oil flask with ether extract-Wt. of the oil flask)×100
basin. The basin was heated in a low flame at the beginning % of Ether extract = ----------------------------------------------------------
till no fumes were given off by the charred mass. It was gm of the substance taken
broken by a glass rod carefully and burnt in a muffle furnace
at 550- 600°C for 4-5 hrs. The muffle was allowed to cool to Determination of crude fibre
150°C. The basin was then cooled in a desiccator and the ash About 2 gm of moisture and fat free sample was weighed and
content was then weighed. The total ash was calculated as transferred to the spout less one litre beaker. Thereafter, 200
follows: ml 1.25% H2SO4 was added. The beaker was placed on hot
% of total ash = weight of the ash ×100 / weight of the sample plate and allowed to reflux for 30 mins, timed from onset of
boiling. The content was shaken after every 5 min. The beaker
Estimation of moisture content was removed from the hot plate and filtered through a muslin
Fresh sample materials were taken in a flat bottom dish and cloth using suction. The residue was washed with hot water
kept overnight in a hot air oven at 100-110°C and weighed. till it was free from acid. The material was transferred to the
The loss in weight was regarded as a measure of moisture same beaker and added 200ml of 1.25% NaOH solution and
content. refluxed for 30 mins. Again filtered and the residue was
washed with hot water till it was free from alkali. The total
Estimation of crude protein (Micro-Kjeldahl Method) residue was transferred to a crucible and placed in hot air
Digestion: About 2gm of sample was taken in a Kjeldahl oven, allowed to dry to a constant weight at 80-110 °C and
flask, 10gms of sodium sulphate and 0.5 gm of copper weighed. The residue was ignited in muffle furnace at 550-
sulphate was added and mixed well. A few glass beads were 600 °C for 2-3 hrs, cooled and weighed again. The loss of
added into the flask to prevent spurting while heating. Then weight due to ignition was the weight of crude fiber.
25 ml of concentrated H2SO4 was added and then heated for
(Wt of the crucible with dry residue -Wt of crucible with ash) ×100
15-20 mins in inclined position. The solution was boiled until
% of Crude Fiber = -----------------------------------------------------------
a greenish colour was obtained. It was allowed to cool. gm of the substance taken

Distillation Procedure for Mineral analysis

About 100 ml of distilled water was added to the Kjeldahl Estimation of Fe, Zn, Mg, Mn, Na, K and Cu
flask, shaken properly and transferred it into a 250 ml For this study, 0.5 gm of powdered dried sample was taken in
volumetric flask. Then the final volume was made up to 250 a crucible and converted to ash in the muffle furnace at 580°C
ml by adding distilled water. In a conical flask, 10-15 ml of for 3 hrs. After cooling in a desiccators 10 ml of concentrated
2% Boric acid was taken and the flask was placed below the Nitric acid, 4 ml of Perchloric acid and 1ml of Sulphuric acid
condenser of the distillation apparatus. Thereafter, 5 ml of was added and digestion at high temperature was carried out
aliquot was transferred to the Micro Kjeldahl steam until the content became clear, then the tube was cooled and
distillation apparatus and added 1 drop of phenolphthalene the solution was transferred quantitatively to 50 ml volumetric
and 10-15 ml 40% NaOH. The distillation was carried out flask and the final volume was adjusted to 50 ml by adding
atleast for 5-10 mins until ammonia was free from aliquot. distilled water. The solution was used for determination of Fe,
Titration: The distillation product was then titrated against Zn, Mg, Mn, Na, K and Cu through the atomic absorption
N/10 H2SO4 spectrometry (AA203D). Calcium and Phosphorous
estimation were done as per method described by Oyodele [14].
Calculation is done as follows:
Results
ml of N/10 H2SO4 used up×250×0.0014×100 Qualitative and Quantitative Phytochemical Analysis of
% of Nitrogen = ------------------------------------------------------ Never Die plant
Volume of aliquot ×gm of the substance taken Qualitative phytochemical compositions of never die plant
(Leaves, Stems and Roots) shows the presence of Alkaloid,
% of crude protein=% Nitrogen ×6.25 Tannin, Saponin, Flavonoid, Terpenoid, Glycoside and
~ 631 ~
Journal of Pharmacognosy and Phytochemistry

Phenols but flavonoids were absent on the roots as shown in Alkaloids, Tannins and Phenols but lowest in flavonoids as
table 1. While quantitative phytochemical composition of shown in Fig. 1
never die plant (Leaves, Stems and Roots) was highest in

Table 1: Qualitative Phytochemical Composition of Never Die Plant

Phytochemical Properties
Plant Samples
Alkaloid Tannin Saponin Flavonoid Terpenoid Glycoside Phenols
Leaves +++ ++ + ++ ++ +++ ++
Stems ++ + ++ + ++ + +
Roots + + + - ++ ++ ++

Fig 1: Quantitative Phytochemical Composition of Never Die plant

Proximate analysis of Never Die plant

Proximate compositions of the leaves, stem and roots of Never Die plant was highest in protein and Moisture contents and lowest
in Lipid and ash contents as shown in Fig. 2.

Fig 2: Proximate Compositions of Never Die Plant

Mineral contents of Never Die Plant Discussion

Mineral contents of Leaves, stems and roots of never die Phytochemical analysis is very useful in the evaluation of
plants shows highest in calcium, sodium and magnesium and some active biological compound of some medicinal plants.
Lowest in zinc, copper and manganese as shown in table 2. The qualitative phytochemical analysis of the roots, leaves
and stems of Bryophyllum pinnatum were carried out and
Table 2: Mineral Content of Never Die Plant alkaloid, glycoside, tannin, phenols, saponin, terpenoid and
Plant Minerals (ppm) flavonoids was present on the roots, stems and leaves of the
Samples Ca Mg Na K Fe Mn Cu Zn PO4 plant samples but flavonoid was absent on the roots.
Leaves 38.47 18.53 29.78 17.17 2.212 0.20 0.12 0.074 0.40 Quantitatively, alkaloids were present in appreciable amount
Stems 28.65 21.46 30.98 18.49 1.489 0.27 0.18 0.055 0.53 (18.72 ± 5.70%). Alkaloids are one of the most efficient
Roots 40.88 36.56 48.72 30.4 2.339 0.24 0.20 0.087 0.66 therapeutically significant bioactive substances in plants. Pure
~ 632 ~
Journal of Pharmacognosy and Phytochemistry

isolated alkaloids and the synthetic derivatives are used as The calcium content (40.88 ± 38.47ppm) was high in the plant
basic medicinal agents because of their analgesic, samples. Calcium helps in the regulation of muscle
antispasmodic and bactericidal properties [15]. Alkaloids tend contraction required by children, infants and foetuses for
to be organic and natural ingredients that have nitrogen, and bones and teeth development [35]. Normal extracellular
are also physiologically active together with sedative and calcium concentration is necessary for blood coagulation and
analgesic roles. They are found in reducing stress and for the integrity, intracellular cement substance [36]. It also
depression symptoms. Alkaloids tend to be poisonous when helps in the development of strong bone and teeth. Potassium
taken in bulk amount due to their stimulatory effects, content (30.4 ± 17.17ppm) was also high in the plant samples,
producing excitation associated with cell and nerve disorders and this is in agreement with many reports that potassium is
[16-17]
. the most abundant mineral in Nigerian agricultural products
[37]
The tannin content (12.40 ± 3.64%) in the plant samples . Potassium helps to maintain body weight and regulate
implies that the leaves, roots and stems of Bryophyllum water and electrolyte balance in the blood and tissues
pinnatum have high astringent properties. Tannins quicken the (National Research Council, NRC), [38]. The concentration of
healing of wounds and inflamed mucous membranes [18]. sodium in the sample was (48.72 ± 29.78 ppm), and supports
Tannins are water soluble phenolic compounds which the claim by the natives that the plant is useful in the
precipitate proteins from aqueous solution. They occur in all treatment of heart related diseases. Excess sodium
vascular plants. Tannins bind to proteins making them bio- consumption leads to hypertension [38].
unavailable [19-21]. The value in this study is in agreement with Zinc is said to be an essential trace element for protein and
the studies associated with other researchers in the same field nucleic acid synthesis and normal body development [39]. Zinc
[22-26]
. Saponin content (2.46 ± 1.20%) suggests the usefulness also stimulates the activity of vitamins, and the formation of
of the plant as a potential fertility agent. The saponin level is red and white blood cells [40]. Zinc plays a role in improving
however low, when compared with the results from other male fertility. The presence of zinc in the plant could mean
works [26-27, 23]. Saponins at low levels < 10% are said to be that the plant can play valuable roles in the management of
safe and non-toxic. Saponins are glycosides containing diabetics which result from insulin malfunction [41-42]. Iron is
polycyclic aglycone moiety of either C27 steroid or C30 said to be an important element in the diet of pregnant
triterpenoids attached to a carbohydrate sugar. High Saponin women, nursing mothers, infants, convalescing patients and
levels have been associated with gastroenteritsis, manifested the elderly to prevent anaemia and other related diseases [43].
by diarrhea and dysentery [28]. The magnesium content of the plants was found to be 36.56 ±
The presence of flavonoids in appreciable amount (2.36 ± 18.53ppm. Magnesium plays fundamental roles in most
0.98%), inferred that the plant samples has the biological reactions involving phosphate transfer. It is believed to be
functions such as anti-oxidation, and protection against essential in the structural stability of nucleic acids. It plays a
allergies, inflammation, free radical, platelet aggregation, significant role in the intestinal absorption of electrolyte in the
microbes, ulcers, hepatoxins, viruses and tumour [29, 18]. body. Its deficiency in man includes severe diarrhoea and
Flavonoids are potent water soluble antioxidants and free persistent migraines [44].
radical scavengers which prevent oxidative cell damage, and Copper level in the plant samples under study ranged from
have strong anticancer and anti-ulcer activity and protection 0.20 to 0.12ppm. Copper is involved in the formation of red
against the different levels of carcinogenesis [29]. Cardiac blood cells and synthesis of haemoglobin. It has a role in
glycosides are important class of naturally occurring drugs energy production, wound healing, skin and hair color as well.
whose actions helps in the treatment of congestive heart Copper is also involved in stimulating body defence system.
failure [30]. In combination with Zinc, it plays a role in superoxide
The moisture content was (3.18 ± 3.13%) indicating that the dismutase activity and the removal of oxygen free radicals [45].
plant is susceptible to spoilage. The ash content (1.87 ± Consumption of manganese-containing foods is believed to
1.81%) is an indication of the level of inorganic elements support the immune system. Manganese regulates blood sugar
such as calcium, zinc, magnesium, copper, and potassium in levels, the production of energy and cell reproduction.
the plant samples. The protein content was (3.79 ± 2.96%) Deficiency in manganese may result in birth defects if an
and readily available as a macronutrient. Protein is an expectant mother does not get enough of this important
essential component of human diet needed for the element [46].
replacement of tissues and for the supply of energy and
adequate amount of required amino acids. Protein deficiency Conclusion
causes growth retardation, muscle wasting, oedema, abnormal The result of this research work shows that the leaves, roots
swelling of the belly and collection of fluids in the body of and stems of Bryophyllum pinnatum contain phytochemicals,
children [31]. High contents of protein in plants are for building proximate and minerals in appreciable quantities and they
and repairing of body tissues, regulation of body processes possess activities like anti-diabetic, anti-ulcer, anthelmintic,
and formation of enzymes, hormones and antibodies that immunosuppressive, hepatoprotective, anti-nociceptive, anti-
enable the body to fight infection [32]. The crude fibre content inflammatory, nephroprotective, antioxidant, analgesic,
(3.10 ± 1.84%) which aid digestion, absorption of water from anticonvulsant, neuropharmacological, antipyretic, and
the body and bulk stool. Fibre softens stool and therefore, antihypertensive. Bryophyllurn pinnatum is also used in the
prevents constipation [33]. The plant may therefore be useful in treatment and prevention of infections. As a rich source of
the control of body weight, blood cholesterol and protection secondary metabolites Bryophyllum pinnatum can be a
against colon cancer. The lipid content (0.73 ± 0.56%) of the potential source of useful drugs. Incorporation of this plant in
plant samples was low, and it can therefore be recommended diet as nutraceuticals is worth recommendation. Moreover,
as part of weight reducing diets. Low lipid foods are said to they are ubiquitous, can be grown or cultivated and is not
reduce the level of cholesterol and obesity [34]. even endangered. We believed that the data provided by us
will be helpful to explore more medicinal plants. Further

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Journal of Pharmacognosy and Phytochemistry

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