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Journal of Mosquito Research, 2013, Vol.3, No.11, 76-81
ISSN 1927-646X
http://jmr.sophiapublisher.com
80
ethyl acetate as solvent (extraction period 72 hour and
the temperature was < 40
℃
). The extract was collected
in a separate beaker.
3.5 Larvicidal bioassay
According to world health organization standard
protocols (WHO, 1981), the larvicidal bioassay with
suitable modification was done. Each of the
previously made concentration of crude extract was
transferred into a sterilized glass petridishes (9 cm
diameter, 150 mm capacity). Twenty larvae of each
larval instar (1
st
, 2
nd
, 3
rd
, and 4
th
) of
Cx.
quinquefasciatus
were separately transferred into
different
petridishes
containing
appropriate
concentration. Larval food (20 mg dried yeast powder)
was added in each Petridish. Mortality rates were
recorded at 24 hr, 48 hr and 72 hr of post exposure.
Dead larvae were recognized when they failed to
move after probing with a needle in the siphon or
cervical region. The experiments were repeated three
times on three different days and maintained at 25~30
℃
and 80%~90% relative humidity.
3.6 Effect on non
-
target organisms
Effect of crude and ethyl acetate extracts were tested
against non-target organisms like Daphnia sp.,
Diplonychus annulatum
(predatory water-bug) and
Chironomus circumdatus
larvae (insect). The
predators were exposed to half lethal concentrations of
crude and solvent extracts at 24 h to observe the
mortality and other abnormalities such as sluggishness
and reduced swimming activity up to 72 h of
exposure.
3.7 Phytochemical analysis of the plant extracts
Phytochemical analysis of the crude extract of seed
coat of
C. sophera
was carried out according to the
methodologies of Harbone (1984) and Stahl (1989).
The phytochemicals included under study were
saponins, terpenoids, alkaloid, steroids, tannin,
flavonoids, cardiac glycosides and free glycoside
bound anthraquinones.
3.8 Statistical analysis
The percentage mortality (%M) was corrected using
Abbott’s formula (1925). Statistical analysis included
the probit analysis (calculating LC
50
and LC
90
values),
regression equations (Y=mortality; X = concentrations)
and regression coefficient values. ANOVA was carried
out to justify the significance between different
variables such as different concentrations, different
instars, hours and mortality rate.
Acknowledgments
The authors acknowledge with thanks the help of Dr. Ambarish
Mukherjee, Professor of Botany, The University of Burdwan,
for identification of the plant. The financial support provided
by the DST, New Delhi, India (D.O. NO.SR/ SO/HS/84/2007
dated 08/02/08) is also acknowledged.
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