The susceptible Cx. quinquefasciatus Sebring strain was used. This strain was originally collected from Sebring, FL, in 1988 (Vanlandingham et al. 2008), and has been colonized at the Centers for Disease Control and Prevention (Fort Collins, CO) since 2004. Adult mosquitoes were maintained at 27.5 °C and 70–80% relative humidity on a photoperiod of 14:10 (L:D) h in environmental chambers. Adult mosquitoes were exposed to compounds at 5–7 d posteclosion.

The compounds tested were thymoquinone, arbutin, thymohydroquinone, hydroquinone, and libocedrol. Hydroquinone and arbutin were purchased from Sigma Aldrich Chemical Co. (St. Louis, MO). Thymoquinone and its derivatives thymohydroquinone and libocedrol were isolated from the steam distillation of incense cedar essential oil. Each compound was serially diluted for a total of five concentrations, producing a range of 3–99% mortality. Twenty-six female Cx. quinquefasciatus mosquitoes, 6–7 d old, were exposed to CO₂ for 5 s then placed on a chill table (BioQuip Products, Rancho Dominguez, CA) maintained between −1 to −7°C. Each female was treated with 0.2 μl of the compound or acetone, which served as the control, on the dorsal thorax using a syringe and repeating dispenser (Hamilton, Reno, NV). Assays were replicated four times. After all individuals were treated for a particular concentration, they were weighed and an average weight per mosquito was calculated. Mosquitoes were maintained on 5% sucrose for 24 h using the same conditions as the colonies. Mortality was recorded after 24 h.

Arbutin is water soluble and did not adhere to the mosquitoes. Several different solvents including acetone, ethanol, and a detergent, tween-20, were combined with the compound. Each female was treated with 0.2 μl of arbutin and the selected solvent and detergent, but no droplet would form that was absorbed by the cuticle. It was determined that the only way to test arbutin was to feed it via a sucrose solution. Sucrose was removed from the mosquitoes 12 h before the assay. Twenty-five mosquitoes were anesthetized with CO₂, so they could be weighed before placing in individual cups. Five different concentrations of arbutin were dissolved in 3 ml 5% sucrose solution and provided to 6–7-d-old females for a 24-h period before mortality was recorded. Dose mortality (LD) data were evaluated using probit analysis generated using SAS software. (Copyright 2002–2010 by SAS Institute Inc., Cary, NC).

The toxicities on adult Cx. quinquefasciatus are presented in Table 1. The LD₅₀ for thymoquinone was 7.51 μg/mg (CI 7.13–7.88) and hydroquinone was 7.79 μg/mg (CI 7.16–8.43). While these values fall within the confidence interval (CI) of each other, the LD₉₅ of these two compounds did not overlap and their slopes are different. Although three of the four related compounds exhibited some level of insecticidal activity, on the basis of both LD₅₀ and LD₉₅ values, only hydroquinone was similar in toxicity to thymoquinone. Hydroquinone was the most toxic metabolite to Cx. quinquefasciatus, followed by thymohydroquinone, libocedrol, and arbutin. Arbutin proved ineffective, as not enough of the compound could be dissolved in the sucrose solution to induce mortality without forming a paste.

The essential oil of incense cedar heartwood, C. decurrens, was previously found to be toxic to three medically important arthropods, Ae. aegypti mosquitoes, Ixodes scapularis (Say) ticks, and Xenopsylla cheopis (Rothchild) fleas (Dolan et al. 2007). Thymoquinone and carvacrol, which are the major components of this oil, are viable compounds for vector control and commercial development (McAllister and Adams 2010). The related compounds of thymoquinone that were evaluated in this study were not as effective as thymoquinone. This suggests that the quinone moiety of thymoquinone is key to its toxicity and derivatization significantly reduces toxicity. Hydroquinone could have the potential for controlling vector mosquitoes, as the LD₅₀ only varied 0.28 μg/mg from thymoquinone.

Thymoquinone has been indicated as having other biological activities of interest for human use. Thymoquinone is the bioactive constituent of the oil of black cumin (Nigella sativa, L.), which has been used since antiquity in the Middle East for treatment of many human diseases (Padhye et al. 2008). This compound has been shown to have antioxidant, anti-inflammatory, and promising anticancer activities in both in vitro and in vivo studies (Gali-Mahtasib et al. 2006, Sutton et al. 2014). It has also been studied as an immune enhancer in Aedes caspius (Pallas) to control disease agents in a novel way (Ahmed et al. 2010). Hydroquinone is commonly used as a skin whitening agent in cosmetics (Tse 2010).

The prospective of using the same application strategies as current commercial insecticides with the compounds evaluated is probably unlikely since they are less toxic than thymoquinone. Many essential oils are known to exhibit repellent, antifeeding, and insecticidal activities against various arthropod species (Isman 2006). The compounds tested here may serve better as natural repellants than insecticides, and should be further evaluated. The benefit of researching and developing natural products based on plant essential oils may result in relatively low-cost production, be environmentally friendly as compared to commercial insecticides, have public favorability, and have different modes of action. However, additional biological activity needs to be considered before insecticide development.