We have been optimizing the method for determination of five (5) pesticides including 2 4-dichlorophenoxyacetic acid (221.04 g/mol), Metalaxyl (279.33 g/mol), Tebuconazole (307.82 g/mol), 4,4'-Dichlorodiphenyltrichloroethane (DDT) 354.48 g·mol−1, and Cypermethrin (416.3 g/mol) using the Agilent 1100 HPLC coupled with the Waters tandem Mass spectrometry with electrospray ionization. We are using masslynx software.

For Mass spectrometry we are using the cone voltage (v) range 10-30; Source temp 80°C; desolvation temp 150°C; desolvation gas flow 650 L/hr; cone gas flow 110 L/hr; and cappilary voltage 3.0 kv.

The stock standard solution was prepared in pure Acetonitrile. For direct infusion, the working standard solution was diluted in 90:10 (HPLC water: acetonitrile). We managed to get the precursor ion and daughter ions of Tebuconazole (308.23 ►56.4, 69.4, 124.7) and Metalaxyl (280.15 ►192, 220.1, 248.1) in positive mode. But for 2 4-dichlorophenoxyacetic acid, 4,4'-Dichlorodiphenyltrichloroethane, and Cypermethrin nothing was observed.

We tried to run LC for two pesticides using a gradient method with a mobile phase composition of HPLC water with 0.1% formic acid (A) and Acetonitrile (B). The gradient was 0.00 min (50% B); 10 min (90% B); 13 min (90% B); 13.10 min (50% B) and 15 min (50% B) at a flow rate of 0.25 mL/min and 400 Bar. The good separation was observed.

For the three pesticides, the literature suggests the use of ammonium acetate. We prepared 2 mM NH4CH2COO in water and used in dilution of the three standards. It worked on Cypermethrin (in positive mode 433.12 ►104.3, 186.6, 268.6) and 2 4-dichlorophenoxyacetic acid (in negative mode 223.02 ►58.4, 94.3, 140.9,) but for 4,4'-Dichlorodiphenyltrichloroethane the precipitate formed. However, we left it.

We tried to run LC using the same gradient method with a mobile phase composition of 2 mM NH4CH2COO HPLC water with 0.1% formic acid (A) and Acetonitrile (B) for all four pesticides. There were no separation observed.

Because the Cypermethrin and 2 4-dichlorophenoxyacetic acid work in the presence of ammonium acetate, we were forced to repeat optimization of Tebuconazole and Metalaxyl with the same dilution solvent. Unfortunately, we didn’t succeed.

We tried to change the cone voltage of 5, to 35 but still no any significant precursor ion observed for the two pesticides.

Please can you assist me on how to optimize it so that the LC method can accommodate all five pesticides?