Extraction, isolation and identification
Stemona roots were washed, dried, sliced into small pieces and ground in powder. Powder (3 kg) was sequentially extracted by maceration with hexane, dichloromethane and methanol, respectively at room temperature for 3 days per extraction. After evaporation of the solvents under reduced pressure, the crude extracts were used for the insecticidal activity to P. xylostella third instar larvae. Ten g portions of the crude extract which exhibited highest mortality of larvae was first fractionated on a silica gel (70-230 mesh) column chromatography and eluted with 20% step gradient of hexane in ethyl acetate and then with methanol. The most active fraction was further isolated by repeated column chromatography and purified by preparative TLC (silicagel 60 F
, Merck) using 20% ethyl acetate in methanol as the developing solution. Identification of the most active compound was carried out by spectroscopic techniques like HR-MS, IR and
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1
NMR.