Identification of compounds in an HPLC fraction from female extracts that elicit mating responses in male screwworm flies,Cochliomyia hominivorax

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Abstract
When hexane extracts of mature screwworm females were chromatographed on a silica gel column, mating stimulant activity was concentrated in a fraction that eluted with hexane-ether (94∶6, v/v). Separation of this fraction with HPLC (acetonitrile-acetone; 60∶40, isocratic) resulted in a chromatogram of some 20 peaks. Only peaks 4–11 elicited mating responses. Peaks 5–10 had most of the activity, with peak 8 producing the highest response. Sixteen compounds were characterized from peak 8 by gas chromatography-mass spectrometry: six unbranched secondary acetates (C31H62O2); seven previously unreported methyl-branched secondary acetates (C32H64O2); one unbranched ketone (C31H62O); and one methyl-branched ketone (C32H64O). The isomeric acetates were not completely resolved from each other by capillary gas chromatography (CGC) on methyl silicone columns. The sixteenth compound was an aldehyde (C30H60O) that was present only in occasional peak 8 preparations. These compounds and several derivatives were characterized by capillary gas chromatography-mass spectrometry (CGC-MS). The position of the acetate group was ascertained by conversion to a keto group or by replacement of the acetate with a methyl group. Pheromone activity was not observed in peaks trapped either from CGC or by recombination of the trapped CGC peaks from HPLC peak 8. This apparent loss of activity from CGC peaks or from TLC cannot currently be explained.