Fig. 1.
Fig. 2.
Fig. 3.
Fig. 4.
Fig. 5.
Optimised tandem mass spectrometry parameters
| Analyte | Retention time (min) | Precursor ion (m/z) | Ion species | Product iona (m/z) | C.E.b (V) | Q1 pre bias (V) | Q3 pre bias (V) | Dwell time (ms) |
|---|---|---|---|---|---|---|---|---|
| Enniatin A | 11.66 | 699.30 | (M+H)+ | 699.3/682.3 | –18 | –32 | –36 | 16 |
| 699.3/210.2 | –31 | –32 | –23 | |||||
| 699.3/100.1 | –55 | –32 | –21 | |||||
| Enniatin A1 | 11.44 | 685.30 | (M+H)+ | 685.3/668.3 | –20 | –32 | –34 | 16 |
| 685.3/210.1 | –29 | –32 | –23 | |||||
| 685.3/100.1 | –55 | –32 | –19 | |||||
| Enniatin B | 11.04 | 657.30 | (M+H)+ | 657.3/640.3 | –18 | –30 | –34 | 16 |
| 657.3/196.2 | –32 | –30 | –21 | |||||
| 657.3/86.1 | –55 | –30 | –18 | |||||
| Enniatin B1 | 11.24 | 671.30 | (M+H)+ | 671.3/654.2 | –18 | –30 | –34 | 16 |
| 671.3/196.1 | –32 | –30 | –22 | |||||
| 671.3/210.1 | –29 | –30 | –23 | |||||
| Beauvericin | 12.26 | 801.30 | (M+NH4)+ | 801/134 | –55 | –22 | –27 | 33 |
| 801/244 | –34 | –22 | –18 | |||||
| 801/784 | –20 | –22 | –30 |
Structural formula of enniatins (ENNs) and beauvericin (BEA)
| Compound | Side chain R1 | Side chain R2 | Side chain R3 |
|---|---|---|---|
| BEA | phenylmethyl | phenylmethyl | phenylmethyl |
| ENN A | sec-butyl | sec-butyl | sec-butyl |
| ENN A1 | sec-butyl | iso-propyl | sec-butyl |
| ENN B | iso-propyl | iso-propyl | iso-propyl |
| ENN B1 | iso-propyl | sec-butyl | iso-propyl |
Validation results for the determination of enniatins and beavericin in fresh and UHT milk
| Parameters | Raw milk | UHT milk | |||||
|---|---|---|---|---|---|---|---|
| Working range | 0.15–50 μg kg–1 | 0.15–50 μg kg–1 | |||||
| LOD (μg kg–1) | ENN A | 0.098 | 0.099 | ||||
| ENN A1 | 0.097 | 0.096 | |||||
| ENN B | 0.098 | 0.098 | |||||
| ENN B1 | 0.088 | 0.089 | |||||
| BEA | 0.095 | 0.095 | |||||
| LOQ (μg kg–1) | ENN A | 0.126 | 0.126 | ||||
| ENN A1 | 0.125 | 0.128 | |||||
| ENN B | 0.128 | 0.124 | |||||
| ENN B1 | 0.129 | 0.130 | |||||
| BEA | 0.101 | 0.099 | |||||
| Spiking level (μg kg–1 | 2.5 | 5.0 | 7.5 | 2.5 | 5.0 | 7.5 | |
| Recoveries (%) | ENN A | 93 | 83 | 84 | 98 | 72 | 77 |
| ENN A1 | 81 | 91 | 88 | 99 | 78 | 76 | |
| ENN B | 87 | 96 | 90 | 81 | 82 | 84 | |
| ENN B1 | 90 | 93 | 79 | 80 | 88 | 85 | |
| BEA | 81 | 92 | 91 | 82 | 79 | 91 | |
| Repeatability (RSDr %)(n = 6) | ENN A | 5.2 | 3.4 | 3.7 | 5.2 | 3.4 | 3.7 |
| ENN A1 | 6.8 | 5.8 | 4.3 | 4.4 | 3.6 | 4.9 | |
| ENN B | 5.1 | 6.5 | 4.6 | 4.9 | 7.3 | 7.1 | |
| ENN B1 | 7.2 | 7.3 | 6.7 | 5.8 | 6.1 | 6.9 | |
| BEA | 5.5 | 8.0 | 7.6 | 3.8 | 5.5 | 7.5 | |
| Within-lab reproducibility (RSDwR %) (n = 18) | ENN A | 10.1 | 14.8 | 11.9 | 13.5 | 7.8 | 16.9 |
| ENN A1 | 12.4 | 17.5 | 16.9 | 11.9 | 15.8 | 6.9 | |
| ENN B | 8.6 | 15.2 | 15.3 | 8.7 | 13.0 | 12.2 | |
| ENN B1 | 12.5 | 14.1 | 12.4 | 9.3 | 11.3 | 8.3 | |
| BEA | 9.8 | 9.7 | 11.0 | 4.9 | 12.0 | 8.9 | |
| Uncertainty expanded (U(y)) (μg kg–1) | ENN A | uc(y) = = 0.55; k = 2, U(y) = 1.10 | uc(y) = 1.10; k = 2, U(y) = 2.20 | ||||
| ENN A1 | uc(y) = 0.66; k = 2, U(y) = 1.32 | uc(y) = 0.78; k = 2, U(y) = 1.56 | |||||
| ENN B | uc(y) = 0.50; k = 2, U(y) = 1.00 | uc(y) = 0.47; k = 2, U(y) = 0.94 | |||||
| ENN B1 | uc(y) = 0.53; k = 2, U(y) = 1.06 | uc(y) = 0.60; k = 2, U(y) = 1.20 | |||||
| BEA | uc(y) = 0.55; k = 2, U(y) = 1.10 | uc(y) = 0.59; k = 2, U(y) = 1.18 | |||||