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International Journal of Horticulture 2014, Vol.4, No.6, 24
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The experimental set up was kept inside the insect
rearing cage for further 30 days for the emergence of the
first filial generation. The percentage number of adult
beetle emergence was calculated according to the
method described by Odeyemi and Daramola (2000).
4.6 Phytochemical screening of
A. boonei
Chemical tests were carried out on the petroleum ether
extract for the qualitative determination of
phytochemical constituents using standard procedures as
described by Harborne (1973), Trease and Evans (1985),
Sofowora (1993).
4.6.1 Alkaloids
A 0.5g sample of the extracts were stirred with 5ml of
1% aqueous hydrochloric acid on a steam bath. 1ml of
the filtrate was treated with a few drops of Dragendorffs
reagent. Turbidity with this reagent will be taken as an
evidence for the presence of alkaloids in the extract. The
extract (1g) were treated with 40% Calcium hydroxide
solution until the extract was distinctly alkaline to litmus
paper and then extracted twice with 10ml portions of
chloroform. The extracts were combined and
concentrated in vacuum to 5ml. The chloroform extract
was then spotted on thin layer plates. Four different
solvent systems (of widely varying polarity) was used to
develop each plant part extract. The presence of alkaloids
in the developed chromatograms was detected by
spraying the chromatograms with freshly prepared
Dragendorffs spraying reagent. A positive reaction on
the chromatogram (indicated by an orange or darker
coloured spot against a pale yellow background) was
used
4
.6.2 Saponins
The ability of saponins to produce frothing in acquenous
solution and to haemolyse red blood cells was used as
screening tests for saponins. For frothning tests, a 0.5 g
sample of the plant extract was shaken with water in a
test tube. Forthning which persists on warming was
taken as preliminary evidence for the presence of
saponins. In order to remove false positive results, the
blood haemolysis test was performed on those extracts
that frothed in water. The extract (0.5 g) was boiled
briefly in 50ml phosphate buffer, pH 7.4 and then
allowed to cool and filtered; 5 ml of the filtrate was
passed (three hours), through an asbestos disc (1.5 mm
thick and 7 mm in diameter), which was previously
soaked with two or three drops of 1% cholesterol in
either and dried. After filtration, the disc was washed
with 0.5 ml of distilled water, dried and boiled in 20 ml
of oxylol for 2 hours to decompose the complex formed
between cholesterol and any saponin in the extract. The
disc was washed in either, dried and placed on a 7%
blood nutrient agar, complete haemolysis of red blood
cells around the disc after 6 hours was be taken as
confirmatory evidence of the presence of saponins.
4.6.3 Tannins
A 5g sample of the plant part extract was stirred with 10
ml of distilled water, filtered, and a few drops of 10%
w/v ferric chloride reagent added to the filtrate. A blue,
black, green or blue-green precipitate was taken as
evidence for the presence of tannins (Trease and Evans,
1985).
4.6.4 Antraquinones
A 5g sample of the plant extract was shaken with 10ml
benzene, filtered and 5 ml of 10% ammonia solution
added to the filtrate. The mixture was shaken and the
presence of a pink, red or violet colour in the ammonia
(lower) phase indicated the presence of free hydroxyl
anthraquinoes. For combined antraquinones, 5g of each
plant part extract was boiled in 10ml aqueous
hydrochloric acid and filtered while hot. The filtrate
were shaken with 5ml of benzene layer, separated and
half of its own volume of 10% (v/v) ammonia solution
added. A pink, red, or violet coloration in the ammonia
phase (lower layer) indicated the presence of
anthraquinone derivatives in the extract (Trease and
Evans, 1985).
4.6.5 Cardiac Glycosides
Legal Test:
The extract (0.5 g) were dissolved in
pyridine and a few drops of sodium nitroprusside
together with a few drops of 20% Na0H were added. A
deep red colour, which faded to brownish, yellow,
indicated the presence of cardenolides.
4.6.6 Flavonoids
Each of the extracts (0.5 g) were dissolved in 5ml
ethanol. This will be shaken and filtered. To 1ml of the
1
100
eggslaid
numberof
Total
emergence
adult
of
number
Total
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Adult
%