Document Type : Research Article
Authors
1
Pesticides Research Department, Iranian Research Institute of Plant Protection, Agricultural Research, Education and Extension Organization, Tehran, Iran.
2
Plant Diseases Research Department, Iranian Research Institute of Plant Protection, Agricultural Research, Education and Extension Organization, Tehran, Iran
Abstract
Introduction
Tomato early blight is an important disease in tomato which is caused by Alternaria alternata, A. tenuissima and A. solani species, and occurs in a wide range of weather conditions all around the world. For the chemical control of this disease, the fungicides Dagonis® (fluxapyroxad 75 g L-1 + difenoconazole 50 g L-1) at a rate of 1200 mL ha-1, Affiance® (tetraconazole 75 g L-1 + azoxystrobin 95 g L-1) at a rate of 600 mL ha-1 and Signum® (boscalid 252 g L-1 + pyraclostrobin 128 g L-1) at rate of 500 g ha-1 were used. In order to evaluate residue levels of these fungicides, experiments were carried out under greenhouse conditions in Alborz province during the years 2020 to 2022.
Materials and Methods
In order to investigate the effect of Affiance®, Dagonis® and Signum® fungicides in controlling tomato early blight disease, experiments with 3 treatments and four replications were conducted in Alborz province under greenhouse conditions in the form of a completely randomized design. Control was considered without any spraying of these fungicides.
In order to measure the residue levels of these fungicides in the treated tomato fruits, samples were collected at 1, 2, 3, 4 and 5 days after spraying according to the Iran’s national standard method no. 8366/2005 entitled “Pesticides- Determination of pesticide residues in crops and livestock- sampling method”. Extraction of pesticides were carried out by Quick, Easy, Cheap, Effective, Rugged, and Safe (QuEChERS) method and pesticide residue levels were measured by Liquid Chromatography Mass/Mass (LC-MS/MS) and the values were compared with the national and international Maximum Residue Limits (MRLs). For preparation and extraction, tomato samples were crushed and homogenized and 15 grams of the homogenized and crushed sample was weighed as a laboratory test sample. By adding 15 ml of acetonitrile containing 1% acetic acid, the overall extraction process was performed. Anhydrous magnesium sulfate, sodium chloride and sodium acetate adsorbents were used to complete the extraction process. By centrifugation, the organic phase was separated from the aqueous tissue and 5 ml of the organic phase obtained from this step was used for the cleanup step. For purification, magnesium sulfate adsorbents were used in order to remove excess water in the medium and PSA (Poly Secondary Amine) in order to remove large molecules, organic acids, proteins and other disturbing co-extractives. Finally, after centrifugation, 1 mL of the resulting organic phase was prepared after filtration for evaporation and then injection into the LC-MS/MS.
Calibration of LC-MS/MS
First, by directly injecting the standard solution (1 µg mL-1) of each of the pesticides alone to the MS detector, the fragmentation voltage of the parent ion (Precursor Ion) and the collision energy for each of the daughter ions (Daughter Ion) or product ions of each compound were optimized. In other words, at this stage, the best conditions for high-sensitivity detection were determined for each of the compounds.
Validation of the Method
According to the Sanco standard, three concentration levels were validated, which were made in acetonitrile solvent and tomato matrix. For this purpose, by diluting the mother solution appropriately, solutions were prepared at three different concentration levels of 0.05, 0.1, and 0.2 mg kg-1 of the mixture of the standards of pesticides under investigation in solvent and tomato matrix. Regarding the linear dynamic range (LDR) for all pesticides the beginning of the range is the same as the limit of quantitation (LOQ). The figures of acceptable merit and recovery in the range of 80.1 to 111% with RSD from 10 to 14.5% indicate the acceptability of the proposed analysis method.
Results and Discussion
The results obtained for the residue levels of Dagonis® fungicide consisting of fluxapyroxad and difenoconazole, showed that according to the MRL of fluxapyroxad (MRL= 0.2 mg kg-1), two days after spraying it was less than the MRL and difenoconazole residue levels was less than the MRL (MRL= 0.6 mg kg-1) one day after spraying. The amount of residue in Affiance® treatment samples, consisting of two fungicides, tetraconazole and azoxystrobin, showed that after 3 days of spraying, the residue of tetraconazole reached the MRL (0.1 mg kg-1) and the residue of the fungicide azoxystrobin was lower than the MRL (3 mg kg-1) one day after spraying. The results of the pesticide residue measured in the samples treated with Signum® consisting of boscalid and pyraclostrobin showed that the pyraclostrobin residue levels were lower that the MRL (1 mg kg-1) two days after spraying this fungicide, and boscalid residue was lower than the MRL (3 mg kg-1) after one day of spraying. In the control samples, the tested fungicides were not detectable.
Conclusions
Therefore, Affiance®, Dagonis® and Signum® fungicides at the rate of 600, 1200 ml and 500 g ha-1, with the pre-harvest intervals of 3, 2 and 2 days respectively, are safe considering the residue levels of them and are recommended to be used to control early blight disease in tomato.
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