Azoxystrobin

The chemical library was donated by the EPA ToxCast Program. For verification and replication experiments, the following chemicals were purchased from Sigma-Aldrich: DL-α tocopherol acetate (T3376), DL-sulforophane (S4441), vincristine sulfate salt (V8879), oxyfluorfen (35031), rotenone (45656), fenamidone (33965), pyraclostrobin (33696), trifloxystrobin (46447), myxothiazol (T5580), pyridaben (46047), azoxystrobin (31697), fluoxastrobin (33797), fenpyroximate (31684) and kresoxim-methyl (37899). Famoxadone was purchased from Chem Service, Inc (N-11943). Topotecan hydrochloride was purchased from Tocris (4562). Paclitaxel was purchased from Fisher Scientific (AC32842). All chemical stocks were prepared in DMSO unless otherwise noted. Vehicle samples were prepared with an equivalent DMSO concentration of ≤0.5% in feeding medium.

The ‘Pathogen Box’ compounds were kindly provided by MMV in 96-well plates containing 10 mM stock solutions dissolved in dimethyl sulfoxide (DMSO). Additional amounts of several compounds were purchased from Sigma Aldrich and dissolved in DMSO (stock concentration given in brackets), including azoxystrobin (31697-100MG; 50 mM), trifloxystrobin (46447-100MG; 50 mM), auranofin (A6733-10MG; 50 mM), buparvaquone (SML1662-25MG; 3 mM), and atovaquone (A7986-10MG; 10 mM). 3-MB-PP1 was purchased from Cayman Chemical (17860; 10 mM). Additional MMV688853 (BKI-1517; 10 mM) was a kind gift from Wes Van Voorhis (University of Washington). Additional MMV024397 was also provided by MMV. The DMSO concentration introduced when using these compounds in assays was < 0.2% (v/v), except MMV688853 when used at the higher concentrations (up to 50 μM) in the Plasmodium assays (up to 0.5% (v/v) DMSO).

Malathion, parathion, sulfotep, fenthion, disulfoton, demeton-O, chloropyrifos and bifenthrin certified analytical standards were purchased from AccuStandard (New Haven, CT, USA). In addition, ethion, carbofuran and azoxystrobin standards were acquired from Sigma-Aldrich (St. Louis, USA) and permethrin acquired from Supelco (St. Louis, USA). All standards presented purity grade of 97% or higher. Acetonitrile (99.99%) (JT Baker, USA), dichloromethane (99.9%) (JT Baker, USA) and methanol (99.9%) (Merck, Darmstadt, Germany) used in this study were spectroscopic and chromatographic grades.
Stock solutions were prepared in methanol at 500 mg L⁻¹ for carbofuran, diazinon, chloropyrifos, ethion, bifenthrin, permethrin and azoxystrobin. The other stock solutions were at 960 mg L⁻¹ (demeton-O), 984 mg L⁻¹ (fenthion), 985 mg L⁻¹ (sulfotep), 990 mg L⁻¹ (molinate), 998 mg L⁻¹ (parathion) and 1000 mg L⁻¹ for Malathion. From the stock solutions, a mix working solution (1000 µg L⁻¹) containing the 13 pesticides was prepared by dilution in methanol. On the other hand, matrix-matching calibration curves composed of seven concentration levels (0.5, 1.5, 2.5, 3.5, 4.5, 5.5, 6.5, and 7.5 µg L⁻¹) were prepared for each compound adding variable volumes of mix working solutions in extracts of PM2.5 blank filters.