Table 3. Phytochemical
constituents of Barleria cristata.L leaf extract
|
S. No |
Phytocostituents |
mg% |
|
1 |
Alkaloids |
0.23±0.092 |
|
2 |
Flavonoids |
4.8±0.104 |
|
3 |
Phenols |
0.104±0.121 |
|
4 |
Tannins |
0.26±0.151 |
|
5 |
Saponins |
3.48±0.527 |
Each value in the table was obtained by calculating the
average of three experiments ± standard
deviation.
Many plant spices have been
investigated in the search of novel antioxidants4. But there is demand still to
find potential antioxidants from the plant sources. With this back ground, the
present study was undertaken to check for the presence of important phytochemicals
such as vitamins, and anti-oxidants in the Berleria cristata plant extract.
MATERIALS AND METHODS:
Collection and identification
of plants:
The plant Barleria cristata L. commonly known as kodilkannu (in
Fresh plant leaves were washed , shade dried and then
homogenized into fine powder by mixer grinder and used for further studies.
Preparation of extract for
qualitative phytochemical analysis:
The powdered plant material was subjected to Soxhelet extraction with petroleum ether, chloroform, 95%
ethanol and distilled water for 18 hours . The condensed extracts were used for
screening of phytochemicals such as alkaloids
(Iodine, Wagner and Dragendorff’s test), flavonoids (Pew’s, Shineda and NaOH test), glycosides (Keller-Killiani,
conc.H2SO4 and Molisch test) Lignins (Labat and Lignin test),
phenols (FeCl3,Leadacetate,Libermann’s tests), saponins
(foam test), sterols (Liberman-Burchard, Salkowski test),
Tannins (FeCl3, leadacetate test), Total carbohydrate (Fehling’s,
Benedict’s test), protein (Biuret, Millon’s test) as reported earlier5,6.
Table 4 . Vitamin composition
of Barleria cristata.L
|
S. No |
Vitamins |
Amount in mg% |
|
1 |
Ascorbic
Acid(C) |
10.7±0.1335 |
|
2 |
Riboflavin(
B1) |
0.22±0.0148 |
|
3 |
Thiamin(
B2) |
0.12±0.120 |
Each value in the table was obtained by calculating the
average of three experiments ± standard
deviation.
Determination of Total Ash:
About 2 g of powdered material was incinerated in a
silica crucible at a temperature not exceeding 450 C until free from
carbon. It was then cooled and weighed. The % w/w of ash was calculated7.
Preparation of alcohol extract:
About 2g of powdered material was soaked with 100 ml of
ethanol in a closed neck flask for 24 hours, shaken frequently to facilitate
the dissolution of alcohol soluble components. 25 ml of the filtrate was
evaporated by drying at 100 0 C to get the known amount of material
which was reconstituted with water after weighing. The % w/w of
ethanol soluble extractive was thus calculated7.
Preparation of water soluble
extract:
The procedure used was same as that of alcohol-soluble
extract but using chloroform and water (1:400 v/v) instead of ethanol7.
Quantitative estimation of phyto-constituents and vitamins:
Presence of secondary metabolites were determined by using standard procedures. Alkaloids by Ikan’s method 8, flavonoids
by-Swain and Hillis method9, phenols by
Bray and Thorpe method10 and tannin by Folin-Denis
method11. The Ascorbic acid
was measured by Barakat et al., method12, Riboflavin and Thiamin were determined by spectro-photometric
method13.
In vitro antioxidant studies:
The free radical
scavenging activity of the plant extract was determined by different in vitro methods viz., DPPH radical by Mensor et al.,14
(2001), Superoxide anion and nitric
oxide radical scavenging activity by the
method reported by Sanchez-Moreno15 (2002). The ability of the
extracts to scavenge hydrogen peroxide (H2O2) was
assessed by the method of Ruch et al.,16(1989).
The percentage of inhibition was calculated using the formula.
(O.D
of control) – (O.D of test) X 100
Scavenging activity =
(% of
inhibition) (O.D of control)
Table 5. Free radical
scavenging activity of 50% ethanolic
extract of Barleria cristata.L at different concentrations
|
Concentration DPA SRA HPA NOA (µg/ml) (%
Inhibition) |
||||
|
100 |
11.8 |
18.4 |
15.2 |
12.3 |
|
200 |
22.7 |
38.0 |
29.28 |
23.6 |
|
300 |
36.6 |
51.4 |
43.7 |
35.8 |
|
400 |
48.26 |
65.48 |
56.6 |
49.2 |
|
500 |
62.0 |
74.6 |
70.3 |
60.5 |
|
IC50(µg/ml) |
6.0 |
9.0 |
7.0 |
6.0 |
DPA – % of DPPH
Radical Scavenging activity
SRA - % of Superoxide Radical Scavenging activity
HPA - % of
Hydrogen Peroxide Radical Scavenging activity
NOA- % of Nitric
oxide radical scavenging activity
RESULTS:
Phytochemical
analysis:
The disease curing properties of medicinal plants are
due to the presence of various secondary metabolites. The leaf extract of Berleria crestata
revealed the presence of
alkaloids, flavonoids, glycosides, phenols, saponins, tannins, carbohydrates and proteins (Table1).
Determination of ash value:
Total ash value of leaf powder was 6.86 mg%. The water
and ethanol insoluble ash were 4.6 mg% and 0.7 mg% respectively (Table 2). This
value is similar to the values reported for
commonly consumed ayurvedic plant Tinospora Tomentosa Miers7.
Determination of ethanol and
water soluble extract:
The yield of water soluble and ethanol extracts were
found to be 36.8 mg % and 1.2 mg % respectively (Table 2).
Quantitative estimation of phyto-constituents and vitamins:
The amount of alkaloids, flavonoids, phenols, tannins
and saponin were determined and found to be 0.23,
4.8, 0.104, 0.26 and 3.48 mg % respectively (Table 3). The plant contains
highest amount of ascorbic acid 10.7 mg %
than other vitamins riboflavin
(0.22mg %) and Thiamin (0.12mg %) respectively (Table 4). Vitamin composition
of this measure was seen in the medicinal plant Aframomum melegueta a Nigerian plant which has antioxidant,
anti-cancer, anti-tumor, anti-viral and anti-inflammotory
activities17.
In vitro antioxidant studies:
The 50% ethanolic extracts of
Barleria cristata exhibited
concentration dependent scavenging activities against DPPH, superoxide anion,
hydrogen peroxide and nitric oxide radicals. The IC50 values were
found to be 6.0, 9.0, 7.0, and 6.0 µg/ml respectively (Table 5).
DISCUSSION:
The Barleria cristata leaves are rich in phytochemicals
such as flavonoids, saponin,
tannins, phenols and alkaloids. The biological function of flavonoids includes
protection against allergies, inflammation, free radicals, platelet aggregation
and viruses18. This may the reason behind the traditional use of Barleria cristata extract
in the treatment of intestinal troubles, rheumatism, tuberculosis and
inflammation.
The presence of phenolic compounds
in the extract indicates that the plant might have antimicrobial activity as
phenolic compounds had been used as standard for the measurement of
bactericidal activities17.
The aim of the
total ash determination was to check the purity and quality of the plant[7.
These determinations indicate the amount of organic and inorganic matter in the
plant sample. The ash content of the leaves (6.86%) is similar to the values
reported for the plant Tinospora Tomentosa found in tropical region of
Barleria cristata
has high quantity of flavonoid and saponin. High tannin
content seen in this plant is responsible for bitter taste, and for astringent
and wound healing properties of the plant (Table). This study also finds that the vitamin C
content is high while vitamin B2, B1 is
low (Table 3). Though the vitamin C content of the plant is low, consumption of
this plant material continuously will provide daily vitamin requirement as
stipulated for healthy adults19. In the composition of its phytoconstituents and vitamin content Barleria
cristata has
shown similarity with the
medicinal plant Aframomum melegueta a Nigerian plant which has antioxidant,
anti-cancer, anti-tumor, anti-viral and anti-inflammatory activities17
(Okwu D E,2005) and hence may possess similar medicinal value.
In living system, free radicals are produced through
metabolic process. The free radicals induced cell damage appears to be the
fundamental cause for number of human disorders. Antioxidant-radical scavengers
are used as preventive agents in the free radical-induced diseases20. (Gyamfi et al.,
1999).
Barleria cristata
showed profound free radical scavenging activity. This beneficial effect of the
plant can be attributed to the presence of secondary metabolites especially due
to the hydroxyl group containing phenolic compounds such as high tannins and flavonoid contents21. In addition to the above
compounds the relatively high vitamin C content of the plant may further confer
it with a very potent antioxidant property22]. Thus the present
study encourages further isolation, purification and elucidation of mechanism
of action of the anti-oxidant compounds from this plant for use as an effective
and valuable medicinal plant.
REFERENCES:
1. Khare C.P., Indian Medicinal Plants: An
Illustrated Dictionary. First edition.
Springer Verlag, 2009.
2. Kumpulainen J.T., Salonen
J,T., 1999. Natural Antioxidants and Anti-carcinogens in nutrition, Health and
disease, The royal Society of chemistry,
3. Cook N. C., Samman,
S. 1996. Flavonoids-
chemistry, metabolism, cardio-protective effects, Source: Journal of
Nutritional Biochemistry, 7: 2, pp.
66-76(11).
4. Koleva II, Van Beek T.
A., Linssen J., P., H., de Groot
A, Evstatieva L.N. 2002. Screening of plant extracts for antioxidant
activity: a comparative study on three testing methods. Phytochemical
Analysis 13: 8-17.
5. Dey P. K., Harborne
J. B., Methods in Plant Biochemistry. Academic Press,
6. Evans W. C., Pharmacognosy. 13thEd, Balliere-Tindall, Londen,1981.
7. Debabrata Devbhuti. 2009. Phytochemical and Acute Toxicity Study on Tinospora Tomentosa Miers. Acta Poloniae
Pharmaceutica-Drug Research, Vol.66 No.1
pp89-92,2009.
8. Ikan R., Natural Products. A laboratory guide,
Academic Press,Londen,1981
9. Swain T and Hillis
W E, S. J. Sci. Food Agric.,1959,10,63.
10. Bray H. G., and Thorpe W. V. 1964. Meth. Biochem. Anal., 1,27.
11. Bajaj, K.L., A.K. Devsharma.
A colorimetric method for the determination of Tannin in tea. Mikochimica Acta (Wien) 1977, II,
249-253.
12. Barakat, M. Z., S. K., Shehab,
N., Darwish and E. I., Zahermy.
1993. Determination of ascorbic acid
from plants. Analyst Biochemistry, 53:
225-245.
13. Okwu, D.E., and O.D. Omodamiro.
2005. Effects of hexane extract and phytochemical
content of Xylopia acthiopica
and Ocimum gratissimum on
the uterus of Guinea pig. Bio-Res.,3:
14. Mensor L.I., Menezes
F.S., Leitao G.G., Reis A.S., Dos
15. Sanchez-Moreno, C., 2002. Review: Methods
used to evaluate the free radical scavenging activity in foods and biological
systems. Food science and technology International, 8, 121-137.
16. Ruch, R. J., Cheng, S. J., and Klainig, J.E., 1989.
Prevention of cytotoxicity and inhibition of
intracellular communication by antioxidant catechins
isolated from Chinese green tea. Carcinogen., 10, 1003-1008.
17. Okwu D. E. 2001. Evaluation of the chemical
composition of indigenous spices and flavouring
Agents. Global J. Pure and Applied Sci., 8:455-459.
18. kwu,D. E. 2004. Phytochemicals
and vitamin content of indigenous spices of south
19.
20. Gyamfi M. A., Yonamine
M., Aniya Y., 1999.
Free-radical scavenging action of medicinal herbs from
21. Das,
N. P., Pereira TA., 1990. Effects
of flavonoids on thermal autooxidation
of palm oil: Structure-activity relationship.
J. American Oil Chemists Society, 67:255-258.
22. Naznin Ara and Nur Hasan.,2009. In
vitro Antioxidant activity of methanolic leaf and
flower extract of Lippia alba. Research Journal of medicinal and
medical sciences. 4(1):107- 110,2009.
Received on
22.09.2009
Accepted on
26.10.2009
© AandV Publication all right reserved
Research
Journal of Pharmacognosy and
Phytochemistry. 1(3): Nov. – Dec. 2009, 209-212