Submitted to: Medical and Veterinary Entomology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 10/15/2008
Publication Date: 3/24/2009
Citation: Akasaka, K., Carlson, D.A., Ohtaka, T., Ohrui, H., Mori, K., Berkebile, D.R. 2009. Determination of HPLC fluorescence analysis of the natural enantiomers of sex pheromones in the New World screwworm fly, Cochliomyia hominivorax. Medical and Veterinary Entomology.23:126-130. Interpretive Summary: Organic chemicals from solvent washes of New World Screwworm female flies were analyzed by a very sensitive new method. The washes contained sex pheromone compounds that have many enantiomers or optically active components, which had never been completely determined due to the tiny quantities present. A scientist from the USDA, ARS, Center for Medical, Agricultural and Veterinary Entomology, Gainesville, Florida, now retired, and another from ARS Midwest Livestock Insects Unit, Lincoln, Nebraska, worked with scientists in Japan to determine the character of the natural materials. A novel method was used that involved attaching a fluorescing chemical molecule to each chemical structure in the extract. Synthesized, authentic versions of two of these sex pheromones were made by cooperators. Only selected natural molecules were detected by this very sensitive method after analysis on a pair of high pressure liquid chromatography column. Three compounds detected were present at 10 to the minus 9 grams per female in colonized flies from Jamaica. The determination of the actual sex pheromones may help solve the problem of suspected poor mating in sterilized flies of this species in Jamaica, and possibly elsewhere.
Technical Abstract: Bioassays of 6 racemic synthesized candidate sex pheromone compounds against male New World Screwworm flies showed that the most potent bioactivity was found with 6-acetoxy-19-methylnonacosane and 7-acetoxy-15-methylnonacosane compared to 4 other isomeric acetoxy nonacosanes. Since all these methyl-branched compounds have 2 asymmetric carbons and 4 possible enantiomers, characterization of the natural enantiomers was essential. All 4 enantiomers for the two most bioactive isomers of the natural sex pheromone had been synthesized for bioassay. Hydrolysis to the alcohol and derivatization of these enantiomers with different fluorescent reagents was followed by column-switched HPLC. The use of two linked, reversed-phase columns of different polarity that were held at subambient temperatures allowed good separation of each enantiomer. This analysis applied to natural material was successful, as (6R,19R)-6-acetoxy-19- methylnonanocosane, and (7R,15R)-, and (7R,15S)-7-acetoxy-15- methylnonanocosane were detected in extracts of recently-colonized female flies in the low femtomole range.