4.4. Larvicidal Screening

Eggs of Aedes aegypti were purchased from Institute of Biotechnology, Vietnam Academy of Science and Technology and maintained at the Laboratory of Department of Pharmacy of Duy Tan University, Da Nang, Vietnam. Adults of Culex quinquefasciatus and Aedes albopictus collected in Hoa Khanh Nam ward, Lien Chieu district, Da Nang city (16°03′14.9″N, 108°09′31.2″E) and were identified by National institute of Malariology, Parasitology and Entomology, Ho Chi Minh City. Adult mosquitoes were maintained in entomological cages (40 × 40 × 40 cm) and fed a 10% sucrose solution and were allowed to blood feed on 1-week-old chicks and mice, respectively. Egg hatchings were induced with tap water. Larvae were reared in plastic trays (24 × 35 × 5 cm). The larvae were fed on Koi fish food. All developmental stages were maintained at 25 ± 2 °C, 65–75% relative humidity and a 12:12 h light:dark cycle at the Laboratory of the Faculty of Environmental and Chemical Engineering of Duy Tan University, Da Nang, Vietnam.

Larvicidal activities of the Cinnamomum essential oils were evaluated according to the protocol Liu and co-workers [66] with slight modifications. For the assay, 150 mL of water that contained 20 larvae (fourth instar) was placed in 250-mL beakers and aliquots of the Cinnamomum essential oils dissolved in EtOH (1% stock solution) were then added. With each experiment, a set of controls using EtOH only (negative control) and permethrin (positive control) were also run for comparison. Mortality was recorded after 24 h and again after 48 h of exposure during which no nutritional supplement was added. The experiments were carried out at 25 ± 2 °C. Each test was conducted with four replicates with five concentrations (100, 50, 25, 12.5 and 6 μg/mL). The data obtained were subjected to log-probit analysis [67] to obtain LC₅₀ values, LC₉₀ values and 95% confidence limits using Minitab^® 19 (Minitab, LLC, State College, PA, USA).