We describe a method using paraffin oil solutions and gas chromatography to measure and correct for differences in volatility among test compounds applied to filter paper and to address the problem of minimizing solvent contributions to EAG responses. To examine the effect of the volatility bias, we evaluated the EAG responses of female fir coneworm, Dioryctria abietivorella, to five volatile plant compounds, using a new method to normalize EAG responses to account for the loss of antennal sensitivity that occurs over time. Stimuli were generated either from equimolar (uncorrected) solutions or from corrected solutions that were adjusted to yield equimolar airborne concentrations in the air puffed over antennae. When uncorrected solutions were tested, the two most volatile compounds, (E)-2-hexenal and (E)-3-hexenyl acetate, elicited significantly larger EAG responses than three terpenes. When corrected concentrations were tested, the ranking of these responses changed: (E)-2-hexenal elicited significantly smaller EAGs than (-)-alpha-pinene, (-)-limonene, and (E)-3-hexenyl acetate. On the other hand, there was no effect on the ranking of EAG responses to the two monoterpenes and a sesquiterpene, (-)-trans-caryophyllene, relative to each other. Normalization of EAG data did not affect the overall results (i.e., stimulus rankings) but did reduce their variance within preparations. The results show that when compounds with widely different volatilities are compared in olfactory bioassays, the concentrations of test solutions should be adjusted to produce emissions with equimolar airborne concentrations.
