Materials and Methods
Extracts
Azadirachta indica leaves were collected from
trees, and were dehydrated in an oven at 40 °C
for 48 to 96 h. Next, they were ground to powder
in a knife mill. To prepare the extract, a sample
of 200 g of powder was divided equally among five
Soxhlet extractors. A filter paper cartridge containing
40 g of powder was placed in each extractor
with 300 mL of solvent. Hexane was the first
solvent used. After 24 h of reflux, the hexane was
removed and dichloromethane added. After 12 h
of reflux, the dichloromethane was removed and
methanol was added. The sample was kept under
reflux with the methanol for 13 h. These reflux
times were chosen to ensure that the extraction
was complete in each case. The completion of the
extraction could be verified by observing that the
solvent became colorless after a sufficiently long
exposure to the sample. This change indicated
that the extraction had reached its limit. The extracts
were concentrated in a rotary evaporator at
40 °C at low pressure using a water column. After
this procedure, the extracts were placed in glass
flasks in a laminar flow cabinet until the solvents
evaporated completely.