Residual analysis of chitosan-based agronanofungicides as a sustainable alternative in oil palm disease management

Oil palm (*Elaeis guineensis*) is a highly productive oilseed crop, but its cultivation is threatened by basal stem rot (BSR) disease, caused by the fungus *Ganoderma boninense*. Conventional fungicides like hexaconazole are effective against BSR, but their environmental impact is significant due to leaching and potential harm to terrestrial and aquatic life. The high residue levels also contribute to soil acidification. Nanoformulations of pesticides offer a potential solution, providing slow release and targeted delivery, thus minimizing environmental impact. This research focuses on evaluating the residual levels of a novel chitosan-hexaconazole nanoparticle formulation, applied via trunk injection, in oil palm tissues and oil. The aim is to assess the efficacy and sustainability of this approach compared to conventional hexaconazole application in managing BSR disease.

Numerous studies highlight the environmental concerns associated with conventional hexaconazole use in oil palm plantations. Research demonstrates the leaching of hexaconazole into soil and water bodies, posing risks to both terrestrial and aquatic ecosystems. The accumulation of hexaconazole residues in soil has also been linked to increased soil acidity. Conversely, research on chitosan nanoparticles as a nanocarrier system for agricultural active ingredients showcases their potential for controlled release, enhanced efficacy, and reduced toxicity. The use of chitosan nanoparticles can improve the solubility, stability, and uptake of fungicides, minimizing environmental impact while enhancing disease control. The QuEChERS method has emerged as a leading technique for rapid and cost-effective pesticide residue analysis in various food and plant matrices.

A chitosan-hexaconazole nanoparticle formulation (18 nm mean diameter) was prepared and applied to oil palm trees using trunk injection at two doses: 4.5 g a.i./palm (single dose) and 9.0 g a.i./palm (double dose). Samples of leaf, stem tissue, crude palm oil (CPO), and crude palm kernel oil (CPKO) were collected at various time points (0, 1, 3, 7, 14, 30, 60, 90, and 120 days after treatment). Hexaconazole residue analysis was performed using a modified QuEChERS method followed by gas chromatography-micro electron capture detection (GC-µECD). The QuEChERS method involved acetonitrile extraction, followed by dispersive solid-phase extraction (d-SPE) cleanup using primary-secondary amine (PSA), graphitized carbon black (GCB), and octadecyl (C18) sorbents. The method was validated for matrix effects, selectivity, sensitivity (LOD and LOQ), and recovery. Matrix-matched calibration curves were constructed, and the dissipation kinetics of hexaconazole residues in leaf and tissue were evaluated using zeroth-order, first-order, and second-order kinetic models. The field trial was conducted in a randomized complete block design (RCBD), with three treatment conditions (control, single dose, double dose) and three replicates per treatment. Statistical analysis was performed using ANOVA and Tukey's test to compare means (p ≤ 0.05).

The analysis revealed that palm oil (both CPO and CPKO) samples were residue-free at all time points. However, significant accumulation of hexaconazole was observed in stem tissue and leaf samples. At the double dose, hexaconazole residues were detectable in leaf samples as early as 6 hours after treatment. The highest concentration in leaf samples was observed at day 30, while in tissue samples, the highest concentration was observed at day 30 for the double dose and day 1 for the single dose. Translocation factor (TF) analysis showed a significant translocation of hexaconazole from stem tissue to the leaves, particularly at the double dose where 100% translocation was observed at day 7. At the single dose, >50% translocation occurred from day 60 onward. The dissipation kinetics of hexaconazole at the double dose in leaf and tissue followed second-order kinetics, with half-lives (t1/2) of 515 and 383 days, respectively. The single dose in tissue followed first-order kinetics with a t1/2 of 147 days. Method validation showed excellent linearity (R² > 0.99) for all calibration curves. The LOD and LOQ values were <2.0 ng/mL for palm oil, <3.0 ng/mL for leaf, and <5.0 ng/mL for tissue. Recovery studies showed recoveries above 100% and RSD values below 3.0%.

The findings demonstrate the potential of chitosan-hexaconazole nanoparticles as a sustainable alternative for managing BSR disease in oil palm. The absence of hexaconazole residues in palm oil is crucial for ensuring food safety, while the high accumulation in stem tissue and leaves suggests improved bioavailability and efficacy in targeting the *G. boninense* fungus. The long half-lives observed indicate a sustained release of the fungicide, potentially leading to prolonged disease control. The superior performance of the nanoparticle formulation compared to conventional hexaconazole application, as indicated by the longer half-life, likely stems from its controlled-release properties and enhanced uptake by plant tissues. The small size of the nanoparticles may facilitate penetration through plant cell walls, enhancing their internalization and systemic mobility within the plant.

This study successfully developed and validated a method for analyzing hexaconazole residues in oil palm samples. The results showed that the chitosan-hexaconazole nanoparticle formulation offers a promising sustainable alternative to conventional hexaconazole for BSR disease management in oil palm. The residue-free palm oil and the high accumulation in stem tissue and leaves, combined with a long half-life, demonstrate the potential for effective disease control with minimal environmental impact. Future research could explore different application methods and formulations to further optimize the efficacy and sustainability of this approach.

The study was conducted in a single location with specific environmental conditions, which might limit the generalizability of the findings. Further studies in different regions with varying environmental factors are needed to validate the results. The study only focused on hexaconazole; further investigation into the residual analysis of other components of the chitosan-hexaconazole nanoparticles would be beneficial. The long half-life of the double dose may necessitate further investigation into potential long-term effects on the oil palm trees and the surrounding environment.