An alternative to synthetic pesticides against two major storage insects (Sitophilus oryzae (L.) and Tribolium castaneum (Herbst)) of rice by Nanoemulsion of eucalyptus oil -

An alternative to synthetic pesticides against two major storage insects (Sitophilus oryzae (L.) and Tribolium castaneum (Herbst)) of rice by Nanoemulsion of eucalyptus oil

To address the health risks associated with chemical pesticides used against storage insects in rice, there is a growing need for safer alternatives. Eucalyptus oil (EO) has emerged as one such alternative. This study focuses on enhancing the effectiveness of EO by preparing nanoemulsions. Various combinations of EO, a surfactant (Tween 80), and water were homogenized to create nanoemulsions. Through centrifugation and various stability tests, the optimal combinations were found to be 1:2 and 1:2.5 (EO:Tween 80) with a 6% EO concentration. The resulting nanoemulsions exhibited droplet sizes of 4.04 nm and 2.27 nm, polydispersity index (PDI) values of 0.37 and 0.77, and zeta potentials of 6.20 mV and 7.69 mV for these two combinations, respectively. The lethal concentration (LC50) of the formulated nanoemulsions against Sitophilus oryzae and Tribolium castaneum pests was significantly lower (0.56 to 0.89 μL/cm2) compared to the LC50 of eucalyptus oil alone (0.795 to 4.178 μL/cm2). These nanoemulsions proved to be more effective than pure eucalyptus oil and could serve as a safe alternative to harmful chemical pesticides for combating stored grain pests.

Material and Methods:

Materials:

Eucalyptus oil (EO) obtained from Nice Chemicals Pvt. Ltd, India

Surfactant (Tween 80, polyoxyethylene sorbitan monooleate) purchased from Merck Specialties Private Limited, India.

Chemical Characterization of Eucalyptus Oil (EO):

Chemical characterization followed the procedure by Thanigaivel et al. (2017).

EO dissolved in high-performance liquid chromatography-grade hexane was injected into Gas Chromatography-Mass Spectrometry (GC-MS).

GC-MS conditions included a capillary column (HP-5MS), helium as the carrier gas, and mass spectra analysis.

Insects Culture:

Tested insects: Sitophilus oryzae (rice weevil) and Tribolium castaneum (rust red flour beetle).

Insects cultured and maintained separately under laboratory conditions at 28 ± 2 °C and 65 ± 5% RH.

Preparation of Nanoemulsion:

Nanoemulsions of eucalyptus oil formulated using tween 80 as the emulsifier.

Different EO concentrations (6%, 8%, and 10%) considered.

EO and tween 80 mixed at various ratios and homogenized for different durations at different rotation speeds.

Formulated emulsions characterized, and pH measured using a pH meter.

Characterization of Nanoemulsion:

Stability Study:

Stress tests conducted, including centrifugation, heating and cooling cycles, and freeze and thaw stress.

Phase separation, froth formation, and other changes noted.

Dynamic Light Scattering (DLS):

Droplet size distribution, polydispersity index (PDI), and zeta potential determined using a particle size analyzer.

PDI values below 0.2 indicate a narrow size distribution.

Bioefficacy Test:

Insecticidal efficiency tested against Sitophilus oryzae and Tribolium castaneum.

EO in acetone applied to petriplates with rice for insect feeding.

Nanoemulsions of EO tested against insects, and mortality recorded 24 hours after treatment.

Statistical Analysis:

Percent mortality calculated from the number of dead insects.

Data analyzed using EPA Probit Analysis Program (Version 1.5) to determine LC50, LC90, LC99 values.

Overview Result and Discussion:

Chemical Constituents of Eucalyptus Oil:

·        Eucalyptus oil major constituents: Eucalyptol (64.80%), α-Pinene (11.17%), β-Pinene (8.19%), γ-Terpinene (5.91%), α-Phellandrene (3.88%), Terpinen-4-ol (0.72%), α-Terpineol (1.01%), and 4-Carene (0.51%).

·        Minor constituents include Carvacrol, α-Terpinene, 1-Epi-alpha-gurjunene, Aromandendrene, Alloaromadendrene, α-Farnesene, γ-Gurjunene, γ-eudesmol, and β-eudesmol.

·        Eucalyptus oil contains terpenoids and phenolic compounds toxic to coleopteran stored pests.

Standardization of Ingredients for Nanoemulsion Formation:

·        Emulsions with 6% eucalyptus oil (EO) concentration were formulated.

·        Optimal nanoemulsion preparation achieved at 15,000 RPM homogenization speed for 10 min.

·        Stable nanoemulsions at EO:tween 80 ratios of 1:2 and 1:2.5 with no phase separation.

Characterization of Nanoemulsions of Eucalyptus Oil:

·        Droplet sizes ranged from 2.27 to 2771.00 nm based on oil concentration and EO:tween 80 ratio.

·        Nanoemulsions with 6% EO exhibited ideal polydispersity index (PDI) values for stability.

·        Higher ζ-potential values (6.20 mV and 7.69 mV) in nanoemulsions indicated increased stability.

·        Nanoemulsions with EO:tween 80 at 1:2 and 1:2.5 exhibited smaller droplet sizes (4.04 nm and 2.27 nm) and higher stability.

Bioefficacy of Nanoemulsion Against Stored Grain Pests:

·        Nanoemulsions tested against Sitophilus oryzae and Tribolium castaneum.

·        Nanoemulsion formulations at 1:2 and 1:2.5 ratios outperformed bulk EO, achieving 100% mortality.

·        LC50 values for nanoemulsions were lower than bulk EO, indicating enhanced efficacy.

·        Nanoemulsion demonstrated 1.4–1.6 times effectiveness against S. oryzae and reduced EO requirement for T. castaneum by 3.5–4.8 times.

·        Downsizing oil particles in nanoemulsion improved contact with insects, enhancing toxicity.

·        Nanoemulsion benefits include water solubility, stability, and uniform dispersion for effective pest management.

·        Overall, nanoemulsion efficiency surpasses that of bulk eucalyptus oil, offering cost-effective pest control.

Conclusion:

Eucalyptus oil nanoemulsion demonstrated superior insecticidal performance. The study highlights that optimal nanoemulsion was achieved with 6% eucalyptus oil (EO) concentration. Conditions for successful nanoemulsion: EO:tween 80 at 1:2.5 ratio, homogenized at 15,000 RPM for 10 min. Improved effectiveness observed against Sitophilus oryzae and Tribolium castaneum. Downsizing natural oil particles enhances efficacy, emphasizing the potential of eco-friendly formulations. The study underscores the significance of utilizing downsized natural oil formulations as effective and environmentally friendly insecticides, offering a safer alternative to chemical pesticides.

Reference:

ADAK, T., BARIK, N., PATIL, N.B., GADRATAGI, B.G., ANNAMALAI, M., MUKHERJEE, A.K. AND RATH, P.C., 2020. Nanoemulsion of eucalyptus oil: An alternative to synthetic pesticides against two major storage insects (Sitophilus oryzae (L.) and Tribolium castaneum (Herbst)) of rice. Industrial crops and products, 143, p.111849.