Myclobutanil

Enantioselective degradation of Myclobutanil and Famoxadone in grape

The enantioselective degradation of myclobutanil and famoxadone enantiomers in grape under open field was investigated in this study. The absolute configuration of myclobutanil and famoxadone enantiomers was determined by the combination of experimental electronic circular dichroism (ECD) and calculated ECD spectra. The enantiomers residues of myclobutanil and famoxadone in grape were measured by sensitive high-performance liquid chromatography/tandem mass spectrometry (HPLC-MS/MS). The linearity, precision, accuracy, matrix effect, and stability were assessed. And the limit of quantification (LOQ) for each enantiomer of myclobutanil and famoxadone in grape was evaluated to be 1.5 and 2 μg kg-1. The myclobutanil and famoxadone showed the enantioselective degradation in grape, and the enantioselectivity of degradation for myclobutanil was more pronounced than that for famoxadone. The half-lives were 13.1 days and 25.7 days for S-(+)-myclobutanil and R-(-)-myclobutanil in grape, separately. The half-life of S-(+)-famoxadone was 31.5 days slightly shorter than that of R-(-)-famoxadone with half-life being 38.5 days in grape. The probable reasons for the enantioselective degradation behavior of these two fungicides were also discussed. The results in the article might provide a reference to better assess the risks of myclobutanil and famoxadone enantiomers in grapes to human and environment. Graphical abstract The enantioselective analysis of myclobutanil and famoxadone in grape.

Myclobutanil enantioselective risk assessment in humans through in vitro CYP450 reactions: Metabolism and inhibition studies

Myclobutanil is a chiral triazole fungicide that is employed worldwide. Although enantiomers have the same physical-chemical properties, they may differ in terms of activity, metabolism, and toxicity. This investigation consisted of in vitro enantioselective metabolism studies that employed a human model to assess the risks of myclobutanil in humans. A LC-MS/MS enantioselective method was developed and validated. The enzymatic kinetic parameters (V_MAX, K_Mapp, and CL_INT) determined for in vitro rac-myclobutanil and S-(+)-myclobutanil metabolism revealed enantioselective differences. Furthermore, human CYP450 enzymes did not metabolize R-(-)-myclobutanil. The predicted in vivo toxicokinetic parameters indicated that S-(+)-myclobutanil may be preferentially eliminated by the liver and suffer the first-pass metabolism effect. However, because CYP450 did not metabolize R-(-)-myclobutanil, this enantiomer could reach the systemic circulation and stay longer in the human body, potentially causing toxic effects. The CYP450 isoforms CYP2C19 and CYP3A4 were involved in rac-myclobutanil and S-(+)-myclobutanil metabolism. Although there were differences in the metabolism of the myclobutanil enantiomers, in vitro inhibition studies did not show significant enantioselective differences. Overall, the present investigation suggested that myclobutanil moderately inhibits CYP2D6 and CYP2C9 in vitro and strongly inhibits CYP3A and CYP2C19 in vitro. These results provide useful scientific information for myclobutanil risk assessment in humans.

Cross-resistance between myclobutanil and tebuconazole and the genetic basis of tebuconazole resistance in Venturia inaequalis

Background: Myclobutanil is one of the most widely used demethylation inhibitor (DMI) fungicides for the management of apple scab, caused by Venturia inaequalis. Strains of V. inaequalis resistant to myclobutanil have been reported across the world. Tebuconazole, another DMI fungicide, has been proposed as an alternative to myclobutanil, and the extent of cross-resistance with myclobutanil therefore needs to be evaluated. The sensitivity to tebuconazole and myclobutanil of a total of 40 isolates was determined. Half the isolates came from an isolated orchard which had never been sprayed with fungicides and half from orchards sprayed regularly with myclobutanil, but still with disease control problems. The progeny of a tebuconazole resistant (R) × sensitive (S) V. inaequalis cross were analyzed in order to improve understanding of the genetic control of tebuconazole sensitivity.
Results: There is cross-resistance between myclobutanil and tebuconazole (r = 0.91; P < 0.001). Sensitivity to tebuconazole of the progeny of a R × S cross varied quantitatively in a pattern which implied at least two gene loci differing between the parental strains. In addition, the asymmetric distribution of the sensitivity in the progeny implied possible epistatic effects.
Conclusion: Resistance to myclobutanil and tebuconazole is strongly correlated. At least two genes are involved in the control of tebuconazole resistance in V. inaequalis.

Myclobutanil-mediated alteration of liver-gut FXR signaling in mice

The effects of exposure to Myclobutanil, a triazole fungicide, on the development and progression of nonalcoholic fatty liver disease (NAFLD) are unclear, but activation of nuclear receptors (NRs) is a known mechanism of azole-induced liver toxicity. Farnesoid X receptor (FXR) is a NR and is highly expressed in the liver and intestine. Activation of FXR tightly regulates bile acid (BA), lipid and glucose homeostasis, and inflammation partly through the induction of fibroblast growth factor 15 (FGF15; human ortholog FGF19). FXR activation is downregulated during NAFLD and agonists are currently being explored as potential therapeutic strategy. In this study, we aimed to clarify the effects of Myclobutanil exposure on FXR activation and NAFLD development. Reporter assay showed Myclobutanil treatment, following FXR activation with potent FXR agonist (GW4064), resulted in a dose-dependent decrease of FXR activity. Furthermore, a 10-day study in male mice demonstrated that cotreatment with Myclobutanil led to an 80% reduction of GW4064-induced ileal expression of Fgf15. In a diet-induced NAFLD study, low-fat diet (LFD) fed mice administered myclobutanil displayed decreased FXR activity in the liver and ileum, while high-fat-high-sugar-diet (HFHSD) fed mice showed an increase in hepatic FXR activity and an induction of target genes regulated by constitutive androstane receptor and/or pregnane X receptor. Our work demonstrates Myclobutanil inhibits FXR activity and modulates FXR activity differentially in mice fed LFD or HFHSD. Our studies suggest the importance of understanding how Myclobutanil could contribute to BA dysregulation in disease states such as NAFLD.

[Determination of myclobutanil enantiomers in wheat and its processed products by ultraperformance liquid chromatography-tandem mass spectrometry based on a chiral stationary phase]

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.