Determination of 11 Triazine Pesticide Residues in Soil by HPLC

Triazine pesticides are organic compounds containing a triazine ring, used primarily as herbicides and insecticides in agriculture. Most triazine herbicides act by inhibiting the D1 protein in photosystem II (PSII), blocking the electron transport chain and preventing the synthesis of ATP and NADPH, which eventually causes weed death from "energy starvation". Newer triazine herbicides, such as indaziflam, target cellulose synthase to inhibit cell wall synthesis and restrict plant growth. Some triazine herbicides irritate skin and eyes; animal studies suggest carcinogenic and teratogenic risks, and long-term exposure may pose health hazards. Due to their long persistence in soil, these compounds can leach into or run off to water bodies, posing risks to aquatic ecosystems.

The analysis was performed on a WAYEAL LC3400 series HPLC system equipped with a UV detector, following the method specified in HJ 1052-2019. The 11 triazine pesticides were well separated, with resolutions exceeding 1.5. Six replicate injections of the 25 mg/L standard solution yielded RSDs for retention time below 0.18% and for peak area below 0.75%, demonstrating good instrument precision. All analytes exhibited linear correlation coefficients greater than 0.9999 within the tested range. When soil samples were spiked with 1.0mg/L of the standard, recoveries ranged from 82.16% to 90.61%, indicating acceptable method performance. No residues of the 11 triazine pesticides were detected in the soil sample tested.

Keywords:triazine pesticides; High Performance Liquid Chromatography (HPLC); UV detection.

1. Instruments and Reagents

1.1 HPLC System Configuration

Table 1 Instrument Configuration

1.2 Reagents and Standards List

Table 2 Reagents and Standards

1.3 Chromatographic Conditions

Table 3 HPLC Operating Conditions

1.4 Materials and Auxiliary Apparatus

Analytical Balance;

Ultrasonic Cleaner;

Vortex Mixer;

Rotary Evaporator;

2.Experimental Methods

2.1 Reagent Preparation

2.1.1 Mixed Working Standard Solution of 11 Triazine Pesticides

Precisely transfer 1.0 mL of the 11 triazine pesticides mixed standard solution (500 mg/L) into a 5 mL volumetric flask, dilute to the mark with acetonitrile, and shake well to obtain the mixed standard stock solution. Then, transfer appropriate volumes of the stock solution and dilute with acetonitrile to prepare a series of mixed working standard solutions at concentrations of 1.0, 10.0, 25.0, 50.0, and 100.0 mg/L.

2.2 Sample Pretreatment 

2.2.1 Test Sample Solution

Carefully transfer 10 g of soil sample into a Soxhlet extraction thimble, and place the thimble into the siphon tube of the Soxhlet extractor. Add 200 mL of acetone-dichloromethane mixed solvent into the bottom flask and perform reflux extraction for 24 h at a reflux rate of 3–4 cycles per hour, then collect the extract.

Line a glass funnel with glass wool or a glass fiber filter membrane, add an appropriate amount of anhydrous sodium sulfate, and filter the extract into a concentrating flask. Rinse the extraction vessel and the funnel with 2–3 mL of acetone-dichloromethane mixed solvent, and combine the rinsates into the concentrating flask.

Concentrate the extract to approximately 0.5 mL, then add about 5 mL of n-hexane and concentrate again to about 1 mL to exchange the solvent to n-hexane for subsequent purification.

Mount the solid-phase extraction (SPE) cartridge onto the SPE manifold. Condition the cartridge sequentially with 5 mL of acetone and 10 mL of n-hexane, and do not allow the cartridge to dry. Before the solvent runs dry, transfer the concentrated extract (approximately 1 mL) onto the cartridge and begin collecting the eluate. Rinse the concentration vessel three times with 3 mL of n-hexane, transfer all rinsates onto the cartridge, and elute with 10 mL of acetone-n-hexane mixed solvent. Collect all eluate.

Concentrate the purified eluate to about 0.5 mL, add approximately 3 mL of acetonitrile, and concentrate again to about 0.5 mL to exchange the solvent to acetonitrile. Dilute to 1.0 mL with acetonitrile for analysis. Blank and spiked samples were prepared following the same procedure.

3.Experimental Results

3.1 System Suitability 

Fig.1 Chromatogram of 11 Triazine Pesticides Mixed Working Standard Solution

Table 4 Test Results of Mixed Working Standard Solution of 11 Triazine Pesticides 

3.2 Precision

Fig.2: Overlaid Chromatograms of Six Consecutive Injections of The 11 Triazine Pesticides Mixed Working Standard Solution (25.0 mg/L)

Table 5 Precision Test Results for Six Consecutive Injections of the 11 Triazine Pesticides Mixed Working Standard Solution (25.0 mg/L).

The precision was evaluated by performing six consecutive injections of the mixed working standard solution at 25.0 mg/L. The RSD values of retention time were below 0.18%, and the RSD values of peak area were below 0.75%, indicating good instrument precision.

3.3 Linear Range

Fig.3 Chromatograms of Selected Triazine Pesticides for Calibration Curve

Table 6 Linearity Test Results for 11 Triazine Pesticides

Linearity results: Within the concentration range of 1.0–100.0 mg/L, the correlation coefficients of the mixed calibration curves for the 11 triazine compounds were all greater than 0.9999, showing satisfactory linearity.

3.4 Limit of Detection

Fig.4 Chromatogram of 11 Triazine Pesticides at 0.12 mg/L

Table 7 Test Results of 11 Triazine Pesticides at 0.12 mg/L

Note: According to the test data of 11 triazine pesticide standards (0.12 mg/L) in the above table, the limits of detection were calculated based on a signal-to-noise ratio of 3. The theoretical LOD values of the 11 triazine pesticides were 0.014, 0.026, 0.013, 0.010, 0.013, 0.012, 0.008, 0.007, 0.008, 0.011 and 0.013 mg/L, respectively.

3.5 Recovery

Fig.5 Overlaid Chromatograms for Recovery Test of 11 Triazine Pesticides

Table 8 Recovery Test Results of 11 Triazine Pesticides

The recovery was evaluated by spiking soil samples with the mixed standard solution at 1.0 mg/L and analyzing them according to the procedure described above. The recoveries of the 11 triazine pesticides ranged from 82.16% to 90.61%, indicating satisfactory recovery performance.

3.6 Sample Analysis

Fig. 6. Chromatogram of the Soil Sample

No residues of the above 11 triazine pesticides were detected in the soil sample.

4.Conclusion

A method for the determination of 11 triazine pesticides in soil was developed following HJ 1052-2019, using a WAYEAL LC3400 series HPLC system with a UV detector. The 11 target analytes were well separated with resolutions greater than 1.5. The instrument precision was satisfactory, with RSDs below 0.18% for retention time and below 0.75% for peak area from six replicate injections of the 25.0 mg/L standard solution. Excellent linearity was achieved over the tested concentration range, with all correlation coefficients exceeding 0.9999. Recoveries at a spiking level of 1.0 mg/L in soil samples ranged from 82.16% to 90.61%. No detectable residues of the 11 triazine pesticides were found in the soil sample analyzed.