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Qualitative phytochemical and physiochemical analysis of Cassia
fistula L. Fruit
Article in Medicinal Plants - International Journal of Phytomedicines and Related Industries · September 2014
DOI: 10.5958/0975-6892.2014.00482.1
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Research Article
Agrawal et al.
Qualitative phytochemical and physiochemical analysis of
Cassia fistula L. fruit
K. Agrawal1, Apurva Joshi3, Shivani Ghildiyal 1, M.K. Gautam 2, Mayank Gangwar 2, R.K. Goel2
and V.K. Joshi 1
1Department of Dravyaguna, Faculty of Ayurveda, Institute of Medical Sciences,
Banaras Hindu University, Varanasi-221 005, India
2Department of Pharmacology, Faculty of Modern Medicine, Institute of Medical Sciences,
Banaras Hindu University, Varanasi-221 005, India
3Department of Pharmaceutics, Indian Institute of Technology, Banaras Hindu University,
Varanasi-221 005, India
ABSTRACT
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The fruit pulp of Cassia fistula Linn. is known to have purgative properties. A specific process, i.e., keeping well
ripe fruits buried within sand for seven days and then drying in sun prior to its medicinal use, has been mentioned
in Charaka Samhita. To explore the effect of above classical processing, a comparative physiochemical and
phytochemical evaluation of classically processed (PD) and non-processed (NPD) fruit pulp of Cassia fistula has
been done together with HPTLC. Results of the study showed presence of alkaloid, glycosides, steroids, triterpenes,
phenolic compounds, tannins, proteins, amino acids and carbohydrats in both types of samples. The extractive
values of PD and NPD were 18.25 % w/v and 15.65% w/v, respectively. The difference in physiochemical analysis
of PD and NPD was also recorded and the HPTLC of PD sample showed an extra peak around Rf 0.76 in comparison
to NPD. These results signify appreciable changes in classically processed sample of Cassia fistula fruit pulp and
provide scientific evidence for the classical processing. [Medicinal Plants 2014; 6(2) : 138-142]
Keywords : Cassia fistula, physiochemical analysis, phytochemical analysis, HPTLC, fruit pulp
INTRODUCTION
Cassia fistula L. is a flowering plant of family
Caesalpiniaceae and native of India, Mauritius, South
Africa, Mexico, Brazil, China, Nepal, West Indies
and East Africa (Chopra, 1956; Kirtikar and Basu,
1975; Chatterjee and Pakrashi, 1992). It is an
ornamental plant due to its beautiful bunches of yellow
flowers. The tree is a medium sized deciduous, 6-9
meters tall with straight trunk and spreading branches.
The plant has got golden yellow coloured flowers and
Corresponding author : V.K. Joshi
e-mail : vkjoshivns@sify.net
Received : January 3, 2014; Accepted : March 24, 2014
doi: 10.5958/j.0975-6892.5.2.009
long stick-like fruit pod with a pungent odour. Cassia
fistula fruit has been advocated as a mild laxative for
children and pregnant women. It is purgative due to
the wax aloin and has been reported to treat many
other intestinal disorders like ulcers (Biswas et al.,
1973; Kirtikar and Basu, 1975; Satyavati and Sharma,
1989). The fruit has also been reported to be a rich
source of potassium, calcium, iron and manganese
(Barthakur et al., 1995). The purgative action of fruit
pulp of C. fistula is due to presence of anthraquinone
(Agrawal et al., 2012). Pharmacological screening of C.
fistula plant was reported to have analgesic, antipyretic
(Patel et al., 1965), sedative (Mazumdar et al., 1998),
anti-hypercholesterolemic (El-Saadany et al., 1991),
anti-inflammatory (Suwal, 1993), hypoglycemic
(Esposito et al., 1991), antipyretic (Kashiwada et al.,
1990), anti-rheumatic (Bodding, 1983), anti-tumor
(Gupta et al., 2000), hepatoprotective (Bhakta et al.,
Medicinal Plants, 6(2) June 2014
Qualitative phytochemical and physiochemical analysis of Cassia fistula L. fruit
139
High performance thin layer chromatography
(HPTLC)
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MATERIALS AND METHODS
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guidelines on quality control method for medicinal
plants (Government of India, 1996; WHO, 1992). The
extracts obtained by cold maceration were subjected to
preliminary phytochemical screening for the presence
of various phytoconstituents like alkaloids, flavonoids,
triterpinoids, carbohydrate, protein, resins, saponins,
steroid, tannin, starch and glycosides by standard
methods (Gautam and Goel, 2012).
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1999), antioxidant (Luximon-Ramma et al., 2002), antifungal and anti-bacterial (Dhar and Qasba, 1984)
properties. Extensive studies have been carried out
during the past few decades on isolation and
characterization of chemical constituents of various
parts of C. fistula. Rhein, glucose, sucrose and fructose
were isolated from the fruit pulp (Lal and Gupta, 1972)
whereas
f istulic
acid,
kaempferol
and
a
leucopelargonidin tetramer having free glycol unit were
isolated from the pods and flowers respectively (Agrawal
et al., 1972). C. fistula pod has been reported to have
very low level of toxicity and no pathogenic effects on
liver, kidney and testis of rat (Akanmu et al., 2004).
The fruit pulp of C. fistula has been well known for
its mild purgative action as explained in Charaka
Samhita (1000 BC to 4th Centuary), the most important
information regarding its use has been described in
Kalpa sthana. It has been advised that full-grown, ripe
fruits possessing good qualities should be collected
and kept buried within sand for a week. Thereafter
fruits should be taken out, dried in sun and then their
pulp is stored for use (Agnivesha, 2007). World Health
Organization has emphasized the need to ensure the
quality of medicinal plant products by using modern
control techniques and applying suitable standards
(Ghildiyal et al., 2012).
The present study has been undertaken to find out
the qualitative phytochemical analysis and
physiochemical changes in both classically processed
(PD) and non processed (NPD) fruit pulp of C. fistula
to validate the fact behind the classical processing.
Plant material
The fruits of Cassia fistula L. were collected from the
campus of Banaras Hindu University during the month
of June 2011. A voucher specimen (CF.12/2011) was
deposited in the Department of Dravaguna, IMS, BHU.
The matured fruits of the plant were divided into two
groups. Group-I was kept buried within sand for a week
(PD) and group-II was not kept within the sand (NPD).
Following the stipulated period both the groups were
dried in sun light, fruit pulp was taken out and were
kept separately in air tight glass jars for further study.
Physiochemical and phytochemical analysis
The values of physiochemical parameters, viz., moisture
content, total ash, foreign matter, acid insoluble ash
and sulphated ash, were determined according to WHO
Medicinal Plants, 6(2) June 2014
Fluorescence analysis
The ground powder was dissolved in the specif ic
reagent, mixed vigorously and filtered through a filter
paper. Finally, it was visualized under long wave UV
(365 nm) as well as in daylight. The color of the
fluorescence was obtained and compared with the
reference standard colour (http://en.wikipedia.org/wiki/
Color_index).
Extractive value
Four hundred mg of classically processed (PD) pulp
and non processed (NPD) pulp samples were weighed
separately, in two glass-stopper conical flasks.
Macerated with 100 ml of distilled water for 6 hours by
shaking frequently and then allowed to stand for 18
hours. Filtered rapidly taking care not to lose any
solvent and 25 ml of the filtrate was transferred to tare
flat-bottomed dishes and evaporated to dryness on a
water bath. Dried at 105°C for 6 hours, cooled in
desiccator for 30 minutes and weighed without delay.
The extractable materials were calculated in mg/g of
air-dried material.
HPTLC Analysis was performed on high performance
silica gel 60 F 254 TLC plates. The plates were precleaned by development to the top with methanol and
dried in fume-hood before use. Sample solutions were
applied to the plate by means of automated spray on
band applicator equipped with 100 ml syringe and
operated with band length 10 mm, application rate 10
s ml–1, table speed 10 mm s -1, and distance from bottom
10 mm. The volume applied for each analysis was 10
ml of the sample. Distance between tracks is 10 mm.
The plates were developed 8 cm beyond the origin
with solution of Toluene: Ethyl acetate: Acetic acid
50:40:10 solvent system as mobile phase in vaporequilibrated twin-trough chamber. After development
the mobile phase was evaporated from the plate by
drying in a fume-hood for 10 min. Then, the CAT-3
software controlling the CAMAG -TLC scanner was
Agrawal et al.
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140
Fig. 1. One extra peak was found to be present around Rf 0.76 in Sample 1 (Pink). A significant change in Rf were obtained.
Compounds present in Sample 1 were found to have more Rf than same Compounds present in Sample 2.
used to scan the developed zones under Long and
Short UV and under white light after derivatization.
RESULTS AND DISCUSSION
The values of physiochemical parameters (Table 1) were
different in both the samples whereas the phytochemical
assessment (Table 2) indicated an identical situation.
In fluorescence analysis study (Table 3) specific colour
appeared with specific reagents. The extractive values
of PD and NPD fruit pulp of Cassia fistula were 18.25
% w/v and 15.65% w/v respectively. HPTLC analysis
showed one extra peak present around Rf 0.76 in PD
Sample (Fig. 1).
Herbal drug which are used in various traditional
medicine needs detailed investigation with
ethanopharmacological approach. The present study
provides information in respect of identif ication,
standardization of herbal drug of C. fistula of Ayurvedic
compendia. Correct identification and quality assurance
of the starting materials is an essential prerequisite to
ensure reproducible quality of herbal medicine which
will contribute to its safety and efficacy (Ghildiyal et
al., 2012). The physicochemical evaluation of the drug
is an important parameter in detecting adulteration or
Table 1. Physiochemical analysis
Parameter
NPD
PD
Total ash
9.425%
12.5%
Acid insoluble ash
4.44%
Water soluble ash
% Extractive value
3.3%
18.25% w/v
4.32%
4%
15.65% w/v
Medicinal Plants, 6(2) June 2014
Qualitative phytochemical and physiochemical analysis of Cassia fistula L. fruit
Table 2. Phytochemical analysis of non processed (NPD)
and processed (PD) samples of fruit pulp of
Cassia fistula
Phytoconstituents
NPD
PD
Alkaloids
+
+
Glycosides
+
+
Flavonoids
-
-
Steroidal/triterpenes
+
+
Phenolic and tannins
+
+
Proteins
+
+
Amino Acids
+
+
Sugars
+
+
+ = Present; – = Absent
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improper handling of drugs. The ash value of the drug
gives an idea of the inorganic composition and other
impurities present in the plant species. Preliminary
phytochemical screening reveals the chemical nature
of the extract and be a sign for the further
pharmacological activity. C. fistula fruit pulp revealed
the presence of alkaloid, glycosides, steroids,
triterpenes, phenolic compounds, tannins, proteins,
amino acids and carbohydrate. Tannins have been
reported to possess antioxidant, wound healing and
antimicrobial activities. Phenolic compounds are
141
commonly known for their anti-oxidant, antiinflammatory and antimicrobial activities, while
triterpenoids have been reported to show
immunomodulatory property (Gautam et al., 2012).
Extractive values are useful for the determination of
exhausted or adulterated drugs and also help in
estimation of specific constituents soluble in particular
solvent. The obtained extractive value of classically
processed (PD) and non processed (NPD) plant material
is 18.25%w/v, 15.65% w/v respectively. The
fluorescence analysis is adequately sensitive and
enables the precise and accurate determination over a
satisfactory concentration range without several timeconsuming dilution steps prior to analysis of
pharmaceutical samples (Pimenta et al., 2006). The
fluorescence colour is specific for each compound. A
non-fluorescent compound may fluorence if mixed with
impurities that are fluorescent. The colour of the powder
of Cassia fistula was observed both under ordinary
light and UV light, which gave specific colour change
present in Table 3. In HPTLC one extra peak was found
to be present around Rf 0.76 in processed sample (Pink).
compounds present in Sample 1 (PD) were found to
have more Rf value than same Compounds present in
Sample 2 (NPD).
The above findings provide scientific evidence for
the classical processing of Cassia fistula fruit pulp for
desirable medicinal effect.
Table 3. Fluorescence powder drug analysis of Cassia fistula L.
S.No.
Powder + Reagent
Fluorescence in daylight
Fluorescence under UV (365nm)
1.
Powder as such
Brown
No Fluorescence (NF)
2.
Powder + 1N NaOH in methanol
Golden red
Dark sea green
3.
Powder + 1N NaOH in water
Peru
Medium Sea Green
4.
Powder + 1N HCl in methanol
Dark golden red
Light green
5.
Powder + 1N HCl in water
Lemon chiffon
Medium aquamarine
6.
Powder + 1N HNO 3 in methanol
Tan
Light green
7.
Powder + 1N HNO 3 in water
Sea shell
NF
8.
Powder + Iodine (5%)
Crimson
NF
9.
Powder + FeCl 3 (5%)
Peru
Dark red
10.
Powder + KOH (50%)
Dark orange
Lime green
11.
Powder + Ammonia (25%)
Golden
Olive green
12.
Powder + Picric acid (saturated)
Yellow
Golden
13.
Powder + Acetic acid
Yellow
Yellow green
Medicinal Plants, 6(2) June 2014
142
Agrawal et al.
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