Waters 2996 array detector

A detailed procedure of extraction and purification of mycotoxins (FBs and BEA) was reported previously [16 ,56 (link)]. The samples before fumonisins (FB₁, FB₂, and FB₃) analysis were derivatized with o-phthalaldehyde (OPA) reagent for 3 min. Methanol: sodium dihydrogen phosphate (0.1 M in water) solution (77:23, v/v) adjusted to pH 3.35 with phosphoric acid was used as the mobile phase with a flow rate of 0.6 mL·min⁻¹. A Waters 2695 apparatus (Waters Division of Millipore, Milford, MA, USA) and a Waters 2475 fluorescence detector (λ_EX = 335 nm and λ_EM = 440 nm) with a C-18 Nova Pak column (3.9 × 150 mm) were used for fumonisins analysis. HPLC analysis of BEA was performed using a Waters 2695 system equipped with a Waters 2996 Array Detector (at 205 nm) with C-18 Nova Pak column (3.9 × 150 mm). Samples were eluted with acetonitrile: water (70:30, v/v) at a constant flow of 1 mL min⁻¹ for 45 min. The limits of detection were 10 and 8 ng g⁻¹ for FBs and BEA, respectively. The obtained positive results (on the basis of retention times) were confirmed by HPLC analysis and compared with the relevant calibration curve (correlation coefficients for FB₁, FB₂, FB_3, and BEA were 0.9967, 0.9983, 0.9966, 0.9991, respectively). Recovery rates for FB₁, FB₂, FB₃, BEA were 93, 96, 87, and 91%, respectively.

The embryo axes (0.5 g) of each variant were homogenized with 75 mL of acetonitrile-methanol-water (16:3:1, v/v/v) and filtered (Whatman no. 4 filter paper). Moniliformin (MON) was extracted and purified according to the methods described by Waśkiewicz et al. [24 (link)]. MON was quantified by high-performance liquid chromatography (HPLC) using a Waters 2695 apparatus (Waters, Milford, MA, USA) with a C-18 Nova Pak column (3.9 × 300 mm, 4 µm) and a Waters 2996 Array Detector (λ_max = 229 nm). Acetonitrile: water (15:85, v/v) buffered with 10 mL of 0.1 M K₂HPO₄ in 40% t-butyl-ammonium hydroxide in 1 L of solvent used as the mobile phase (flow rate  =  0.6 mL min⁻¹) [68 (link)]. Quantification of MON was performed by measuring the peak areas at the MON retention time according to the relevant calibration curve (the correlation coefficient for MON was 0.9990). The recovery for MON was 90%, the limit of detection was 1.0 ng g^−1, and the standard deviation was below 7%. All samples were injected in triplicate.