Determination of the Relative and Absolute Configu

Authors: Joshua Rodstein Jocelyn G Millar James D Barbour J Steven McElfresh Ian M Wright Karen S Barbour Ann M Ray Lawrence M Hanks

Publish Date: 2010/12/03

Volume: 37, Issue: 1, Pages: 114-124

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Abstract

We previously identified the basic structure of the femaleproduced sex attractant pheromone of the cerambycid beetle Prionus californicus Motschulsky Cerambycidae Prioninae as 35dimethyldodecanoic acid A synthesized mixture of the four stereoisomers of 35dimethyldodecanoic acid was highly attractive to male beetles Here we describe stereoselective syntheses of three of the four possible stereoisomers and the results of laboratory and field bioassays showing that male beetles are attracted specifically to 3R5S35dimethyldodecanoic acid but not to its enantiomer 3S5R35dimethyldodecanoic acid indicating that the 3R5Senantiomer is the active pheromone component The diastereomeric 3R5R and 3S5Senantiomers were excluded from consideration because their gas chromatographic retention times were different from that of the insectproduced compound The mixture of the four stereoisomers of 35dimethyldodecanoic acid was as attractive to male P californicus as the 3R5Senantiomer indicating that none of the other three stereoisomers inhibited responses to the active enantiomer Beetles responded to as little as 10 ng and 10 μg of synthetic 35dimethyldodecanoic acid in laboratory and field studies respectively Field studies indicated that capture rate did not increase with dosages of 35dimethyldodecanoic acid greater than 100 μg In field bioassays males of a congeneric species P lecontei Lameere were captured in southern California but not in IdahoVolatile maleproduced sex or aggregation pheromones have been reported for an increasing number of cerambycid beetle species Silk et al 2007 Lacey et al 2009 Millar et al 2009 and references therein Ray et al 2009a b most of which are members of the subfamilies Aseminae and Cerambycinae Monné and Hovore 2005 Femaleproduced pheromones also have been identified for two species that formerly were placed in the Cerambycidae Migdolus fryanus Westwood and Verperus xatarti Dufour Leal et al 1994 Boyer et al 1997 but whose familylevel taxonomy is uncertain Napp 1994 Bense 1995 Dong and Yang 2003 Many cerambycid species in the subfamily Prioninae have morphological and behavioral traits that are associated with production of pheromones by females in other types of insects Thornhill and Alcock 1983 Gemeno and Schal 2003 including more elaborate antennal structures in males than females and a more sedentary nature in females than males Summerland 1932 Benham and Farrar 1976 Santos Ferreira 1980 Hovore 1981 Moreover males of some prionine species display behaviors indicative of responses to volatile pheromones produced by females Rotrou 1936 Edwards 1961 Benham and Farrar 1976 Gwynne and Hostetler 1978The prionine species Prionus californicus Motschulsky is broadly distributed in western North America and is an economically important pest of hop Humulus lupulus L Urticales Cannabaceae in the northwestern United States Bishop et al 1984 Adult P californicus are active during summer and early fall and are crepuscular Linsley 1962 They are among the largest cerambycids in North America ranging in body length from 25 to 45 cm volumes indexed in Linsley and Chemsak 1997 Adult females oviposit in soil near the base of living woody plants larvae feed on roots and rhizomes requiring 3–5 yr to complete development Linsley 1962We previously reported that female P californicus produce a volatile sex pheromone associated with an eversible structure on the ovipositor and that males are strongly attracted to this pheromone Cervantes et al 2006 Barbour et al 2007 The basic structure of the pheromone was identified as 35dimethyldodecanoic acid by gas chromatographic GC retention time and mass spectral matches with those of one of the two peaks in a synthetic standard composed of an approximately equal mixture of all four stereoisomers of this compound Rodstein et al 2009 In the work reported here we determined that the biologically active stereoisomer is 3R5Sdimethyldodecanoic acid and confirmed the activity of the synthetic pheromone in laboratory and field bioassays We also showed that the mixture of stereoisomers is as active as the insectproduced enantiomer indicating that the unnatural stereoisomers do not antagonize attraction to the natural enantiomerLarvae of P californicus were collected in fall of 2007 from commercial hop yards in southwestern Idaho and reared to adulthood in a field plot at the Southwest Idaho Research and Extension Center Canyon Co ID USA Larvae were reared individually in 19l plastic buckets buried in the ground to within ~10 cm of their tops Each bucket had two 075 cm holes in the bottom for drainage and buckets were filled with soil to ~10 cm from the top We collected late stage larvae 5–75 cm long from uprooted hop crowns and buried each larva ~10 cm deep in the soil in a bucket with a section of fresh hop rhizome for foodWe cut back hop plants growing from rhizomes in the buckets in early spring 2008 and we fitted buckets with lids that had the centers replaced with aluminum screen Each bucket was dripirrigated 8 l/hr for 10 min every other day Adult beetles emerged in buckets in July 2008 and were collected and held individually in the laboratory in ventilated glass jars ~500–1000 ml 22°C 168 hr LD Males and females were unmated and held in separate growth chambers Beetles used in bioassays were 1–14dold vigorous and apparently healthyTetrahydrofuran was dried and purified by distillation from sodiumbenzophenone ketyl under argon Solutions of crude products were dried over anhydrous Na2SO4 and concentrated by rotary evaporation under reduced pressure unless specified otherwise Crude products were purified by flash or vacuum flash chromatography on 230–400mesh silica gel 1H and 13CNMR spectra were recorded on a Varian INOVA400 spectrometer Palo Alto CA USA 400 and 1005 MHz respectively as CDCl3 solutions Integer resolution mass spectra were obtained with a HewlettPackard HP 6890 GC Avondale PA interfaced to an HP 5973 mass spectrometer MS in EI mode 70 eV with helium carrier gas The GC was equipped with a DB5MS column 25 m × 020 mm ID × 025 μ film Resolutions of enantiomers were attempted on a CyclodexB column 30 m × 03 mm ID JW Scientific Folsom CA USA using isothermal conditions or temperature programs of 2–4°C per min Optical rotations were measured in CH2Cl2 on a Perkin Elmer 241 polarimeter Na and Hg lamps Perkin Elmer Waltham MA USA in a 1 ml waterjacketed cell 10 cm path length at 24°C All rotations were averaged over three or four measurements IR spectra were taken as films on NaCl plates on a Bruker Equinox 55 FTIR instrument Freemont CA USASynthesis of 3R5S+35dimethyldodecanoic acid a acetyl chloride MeOH b MeMgBr CuI STolBINAP tbutylmethyl ether c DIBALH hexane d methyl triphenylphosphoranylidene acetate THF e MeMgBr CuI RTolBINAP tbutylmethyl ether f KOH MeOH/H2O

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