Residual fate of fenazaquin (10EC) in apple fruit and soil

2.1 Field trial

The experiment was carried out on a 20 year old commercial orchard at Tel Bal area of Srinagar, Jammu & Kashmir, where apple variety “Red Delicious” was selected for testing fenazaquin 10 EC (Magister) for dissipation of residues at the recommended (0.004%) and double the recommended (0.008%) application rates. The trees were planted in contour system with plant-to-plant distance of 8.0 m. The trees were at a good bearing stage producing good quality fruit. The experiment was conducted in a single tree plot replicated five times in a randomized block layout. A total of fifteen trees including control (water spray) were selected and marked for the study. The trees were sprayed with fenazaquin one month prior to harvest and a complete coverage of plants was assured.

2.2 Meterological data

The weather conditions play an important role in the dissipation of pesticide residues. Meteorological data including temperature, rainfall and humidity during the period of field experiments were obtained from Meteorology Section of Division of Agronomy, SKUAST-Kashmir. The average maximum and minimum daily air temperatures (^oC) recorded during sampling period ranged from 17.5 to 33.0 and 8.0 to 12.6, respectively. The minimum and maximum relative humidity in the range of 36–97 per cent and the rainfall below 2 mm were recorded during the experimental period.

2.3 Field sampling

Samples of apple from the sprayed trees were obtained randomly from each replication for the estimation of pesticides. Samples were collected at 0, 3, 7, 10, 15, 20 days and at harvest. Zero day samples consisted of fruits collected within one hour of spraying when the spray fluid had dried up. Harvest samples were taken 30 days after the application of treatment. The samples were collected in polythene bags (I kg capacity) with dry ice and taken to the laboratory within 1/2h from the time of collection. The samples were stored in the deep freezer (-20 °C) in order to avoid any degradation of residues between sampling and analysis.

2.4 Extraction and clean up

Apple fruits (1 kg) were chopped (with peel and pulp intact) on a cutting board and put in a blender and blended at 1000 rpm. The samples were processed for fenazaquin residues as described by Kadenczki et al. (1992). The representative 50g of finely homogenized sample was extracted with 200 ml acetone and hexane (1:1 V/V) and shaken for two hours at 80 cycles per minute in a horizontal shaker. The extracted samples were filtered through glass funnel using glass wool. The sample was transferred to the separating funnel and partitioned twice with 75 ml ethyl acetate after dilution with 10 per cent NaCl solution. The organic phase was passed through anhydrous Na₂SO₄ and concentrated in a rotary evaporator to reduce the volume up to 10 ml.

Glass column (60 cm long and 22 mm diameter) was packed compactly with activated charcoal and activated florisil (1:5 W/W). The column was pre-wetted with 10 ml hexane. The concentrated extract was loaded in the column and eluted with 125 ml hexane and ethyl acetate mixture (1:1 V/V). The eluate was concentrated on vacuum evaporator. The final volume was made up to 5 ml in n-hexane. The final extract was analysed on a Varian 450 (Walnut Creek, CA, USA) gas chromatograph (GLC) equipped with an Thermionic Specific detector (TSD Ni63), capillary column CP SIL 8 CB (25 m × 0.25 µm ID × 0.25 µm film thickness of 5% diphenyl and 95% dimethylpolysiloxane). The operating parameters of the instrument were as follows:

• Injector temperature : 240 °C

• Detector temperature : 250 °C

• Oven temperature : 170 ⁰C for 2 min

• 06° C/min upto 230° C and held for 5 min

• 10° C/min upto 260° C and held for 5 min

• 10° C/min upto 270° C and held for 5 min

Nitrogen was used as carrier gas with flow of 1 ml min − 1 through the column and detector make-up of 30 ml min − 1. Under these operating conditions, the retention time of fenazaquin was 9.43 min. Total run time was 31 min. Galaxy chromatography data system version 1.9.302.530 was used for the instrument control and data processing.

2.5 Preparation of standard

Analytical grade reference standard procured from Dr. Ehrenstorfer GmbH, Germany was used for standard curve preparation. A stock solution of 1000 μg g − 1 was prepared from which different concentrations ranging between 0.01 and 1.0 μg g − 1 were obtained for the preparation of standard curve. The data was subjected to regression analysis from which it is evident that the pesticide followed a linear relationship showing corresponding increase in respective pesticide concentrations with coefficient of determination (R²) of 0.9988.

2.6 Recovery

Before analyzing actual sample of fruits, the efficiency of the method was evaluated in recovery experiments by spiking untreated samples of fruits (collected from control plots) with different pesticides. A 50 g well homogenized sample of fenazaquin was spiked with known amount of standard pesticides at three concentrations each replicated thrice. The fortified samples were extracted and cleaned with the method followed for analyzing actual samples. The recoveries for the three levels fell within the acceptable tolerance of 70 to 120 per cent range (SANCO/12571/2014) indicating good performance of extraction, clean up and chromatographic parameters for residue determination in fruits (Table 1). The relative standard deviations (RSDs) were less than 20 per cent for all the levels. In the present study, fenazaquin was used against European red mite (Tetranychus ulmi koch) and analysed for residues. Crop yield and crop health were satisfactory as other pesticides were also used for the comprehensive protection of the apple crop.

2.7 Analysis of data

The data was subjected to statistical analysis as per Hoskin (1961): $$\text{Residue}\left(\mathit{\mu }\text{g g}-\mathbf{1}\right)=\frac{\text{Standard Injected}\left(\text{ng}\right)}{\text{Area of standard}\left(\mu \text{V}.\text{Min}\right)}\times \frac{\text{Area of sample}\left(\mu \text{V}.\text{Min}\right)}{\text{Sample injected}\left(\mu \text{l}\right)}\times \frac{\text{Sample volume}\left(\text{ml}\right)}{\text{Sample weight}\left(\text{g}\right)}$$

The half-life value of insecticides as indices of the rates of residue dissipation was calculated as per Hoskins (1961): $${\mathit{T}}_{\mathbf{1}/\mathbf{2}}=\frac{0.301}{b}$$

b = slope of regression equation

Withholding period (T_tol) based on the prescribed maximum residue limits (MRLs) by European Union was worked out as indices of the safety to consumers. $${\mathit{T}}_{\mathit{tol}}=\frac{\text{a}-log\left({10}^3\text{x}\text{MRL}\right)}{\text{B}}$$

2.8 Determination of residues of fenazaquin in soil under treated apple tree canopy

In this study, the soil samples (1 kg each) were collected in labeled wide mouth amber glass bottle using stainless steel auger (15 cm deep and 3–5 cm diameter) around the trees along the four directions that were sprayed with various pesticides for study of dissipation at various intervals. 5 samples were thoroughly mixed to ensure that the soil collected was truly representative of each treatment. Soil samples were subsequently taken to the laboratory where they were air dried at room temperature, powdered in a pestle and mortar, sieved through a 2 mm sieve and stored at −20 0C until further chemical processing. The sampling was carried out at 0, 3, 7, 10, 15, 20 days after treatment and at harvesting of the fruit.

2.9 Extraction and clean up of soil sample

The extraction and clean up method for analysis of fenazaquin from soil samples was carried out as per Kadenczki et al. (1992). The soil sample was extracted with 150 ml acetone and hexane (1:1V/V). The extract was partitioned with ethyl acetate after dilution with 10 per cent NaCl solution and concentrated in a rotary evaporator. A final analysis was carried out on GLC/NPD.

2.10 Recovery

GC method was used for analysis of fenazaquin residues in soil. Before analyzing the actual samples of soil, the efficiency of the method was evaluated in recovery experiments by spiking untreated samples (collected from control plots) with fenazaquin. A 50 g well ground soil sample of, fenazaquin was spiked with known amount of fenazaquin at three levels. For spiking level, three replicated samples were extracted with respective solvents and cleaned up on florisil column and analyzed by GC. $$\text{Residue}\left(\text{ppm}\right)=\frac{\text{RA}\times \text{TS}}{\text{SA}\times \text{SW}}$$

Where,

RA = Residue in aliquat (µg) estimated by analytical procedure

TS = Total solvent (ml) added per sample

SA = Size of aliquat (ml)

SW = Sample weight (g)

Half life values (T_1/2) corresponding to rate of dissipation were calculated by the method of Hoskins (1961). $${\text{T}}_{1/2}=\text{log}2/\text{b}$$ where b = slope of regression equation

The waiting period (T tol) required to be elapsed for the pesticide deposits to reach the maximum residue limit required for the safe consumption of the fruit after pesticide application was also worked out by the method of Hoskins (1961).