Abstract:

A valid method based on ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) with a chiral stationary phase was established for the determination of myclobutanil enantiomer residue in wheat grain and its processed products (flour, bran, pasta, steamed bun, noodle, and cooking water). The wheat grain and processed product samples were extracted with acetonitrile and purified with primary secondary amine (PSA) and C18. The enantiomers of myclobutanil were separated by Chiral column Lux Cellulose-1 (150 mm×2.0 mm, 3 μm, Phenomenex). The column temperature, sample volume injected, and flow rate were 30 ℃, 5 μL, and 0.25 mL/min, respectively. The mobile phase consisted of phase A (25%), water with 0.1% formic acid and 4 mM ammonium acetate, and phase B (75%), methanol with 0.1% formic acid and 4 mM ammonium acetate. A Waters Xevo TQ-S Micro MS/MS system (Waters, USA) was used for mass spectrometric analysis. An electrospray ionization (ESI) source operating in the positive ionization mode. MS analyses were performed in the multiple reaction monitoring (MRM) mode. The qualitative ions of myclobutanil were m/z 288.9/69.9 and 288.9/124.9, and the quantitative ion of myclobutanil was m/z 288.9/69.9. The source voltage was 3000 V, and the desolvation temperature was 400 ℃. The desolvation gas flow was 800 L/h, and the source temperature was 150 ℃. The matrix effect of wheat grains and their processed products on the determination of myclobutanil enantiomers by UPLC-MS/MS was investigated. S-(+)-myclobutanil and R-(-)-myclobutanil had a mid signal suppression effect on wheat grain, bran, pasta, steamed bun, and noodle, while S-(+)-myclobutanil and R-(-)-myclobutanil had a mid signal enhancement effect on flour and cooking water. Finally, the matrix-matched calibration method was effective in all matrices and was selected for the quantification of the myclobutanil enantiomer residue in the samples. The results showed that the two enantiomers of myclobutanil were well separated by this method. The first and second eluted enantiomers were S-(+)-myclobutanil and R-(-)-myclobutanil, respectively, with the corresponding retention times being 4.34 min and 5.13 min. The limits of detection (LOD) and limits of quantification (LOQ) of S-(+)-myclobutanil and R-(-)-myclobutanil in wheat and its processed products were 0.2 μg/kg and 0.5 μg/kg, respectively. In the linear range of 0.5-25 μg/L, the peak areas of the myclobutanil enantiomers showed a good linear relationship with the concentration, and the R² values were all greater than 0.99. At fortification levels of 5, 50, and 100 μg/kg (enantiomer concentration), the average recoveries of S-(+)-myclobutanil in wheat grain and its processed products ranged from 82% to 110%, with RSDs between 0.9% and 6.8%. The average recoveries of R-(-)-myclobutanil in wheat grain and its processed products ranged from 80% to 109%, with RSDs between 0.9% and 6.8%. This method fulfils the requirements for pesticide residue analysis. The established method was applied to analyze five flour samples, two noodle samples, and two steamed bread samples. The results showed that S-(+)-myclobutanil and R-(-)-myclobutanil enantiomers were not detected in the samples. In this study, methods for the enantiomeric separation and residue analysis of myclobutanil in wheat were evaluated at the enantiomeric level, which enriched the methods of enantiomeric separation and residue analysis of chiral pesticide myclobutanil enantiomers in raw agricultural product (wheat grain) and its processed foods. This method is effective for the residue analysis of chiral pesticide myclobutanil enantiomers in raw agricultural commodities and its processed products.

Key words: chiral stationary phase, ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS), myclobutanil, enantiomers, wheat