2.6.2. Malathion metabolism
Batches of 25 mosquitoes (about 40–60 mg wet weight) were
homogenized in 0.5 ml of 25 mMTris–HCl buffer (pH 7.5) and centrifuged
at 13,000g for 5 min. Supernatant was incubated with
300 lM malathion for 2 h at room temperature. The mixture was
then extracted twice with 0.5 mL acidified chloroform. The chloroform
extract was dried under a current of air, re-dissolved in 30 lL
acidified chloroform and loaded onto a thin layer chromatography
plate. After running with n-hexane: diethyl ether (1:3) the plate
was sprayed with 0.5% (w/v) 2,6-dibromoquinone 4-chloromide
in cyclohexane and left at 100 C for 2 h to visualize malathion
and its metabolic products. Tris buffer (0.5 mL), incubated with 1
lL of malathion and NaOH solution (10 lL of 300 lM Malathion
+ 10 lL of 300 lM NaOH and heated at 80 C) was run as a
positive control. Tris buffer (0.5 mL), incubated with 1 lL of
300 lM malathion, was run as a negative control.