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Development, in-house validation and application of a method using high-performance liquid chromatography with fluorescence detection (HPLC-FLD) for the quantification of 12 ergot alkaloids in compound feeds

Ewelina Kowalczyk
Ewelina Kowalczyk
 und 
Krzysztof Kwiatek
Krzysztof Kwiatek
   | 19. Dez. 2023
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Online veröffentlicht: 19. Dez. 2023
Seitenbereich: 603 - 610
Eingereicht: 17. Juli 2023
Akzeptiert: 05. Dez. 2023
DOI: https://doi.org/10.2478/jvetres-2023-0070
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© 2023 Ewelina Kowalczyk et al., published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

Fig. 1.

Example chromatogram obtained in the course of method optimisation. A 0.2 g mass of primary-secondary amine and 0.01 g of activated carbon were used for clean-up. LU – luminescence unit
Example chromatogram obtained in the course of method optimisation. A 0.2 g mass of primary-secondary amine and 0.01 g of activated carbon were used for clean-up. LU – luminescence unit

Fig. 2.

Chromatograms of A – blank feed sample; B – blank feed spiked at the concentration of 10 μg kg−1 1 – ergometrine; 2 – ergometrinine; 3 – ergosine; 4 – ergotamine; 5 – ergocornine; 6 – ergocryptine; 7 – ergocristine; 8 – ergosinine; 9 – ergotaminine; 10 – ergocorninine; 11 – ergocryptinine; 12 - ergocristinine
Chromatograms of A – blank feed sample; B – blank feed spiked at the concentration of 10 μg kg−1 1 – ergometrine; 2 – ergometrinine; 3 – ergosine; 4 – ergotamine; 5 – ergocornine; 6 – ergocryptine; 7 – ergocristine; 8 – ergosinine; 9 – ergotaminine; 10 – ergocorninine; 11 – ergocryptinine; 12 - ergocristinine

Validation results for determination coefficient (R2), matrix effect, limit of detection (LOD) and limit of quantification (LOQ) and uncertainty of analysed ergot alkaloids

  Linearity Matrix effect (%) LOD (μg kg−1) LOQ (μg kg−1) Uncertainty (%) (at concentration:)
R2   10 μg kg−1 250 μg kg −1 1,000 μg kg −1
ergocristine 0.9992 y = 3.313x−14.6219 65.2 2.2 3.8 17.1 15.4 16.8
ergocristinine 0.9996 y = 3.8130x−27.8254 97.1 1.0 2.0 24.9 4.5 7.3
ergotamine 0.9998 y = 25522x−22.9181 132.8 2.2 4.6 30.6 16.2 11.6
ergotaminine 0.9998 y = 8.3582x−45.1085 68.7 2.6 4.3 36.3 11.4 7.0
ergocryptine 0.9984 y = 1.5186x−8.3205 85.4 2.1 5.0 20.9 10.3 12.9
ergocryptinine 0.9997 y = 4.2522x−29.9617 94.1 1.6 3.5 26.3 4.9 10.0
ergometrine 0.9903 y = 3.3136x−14.6219 42.3 2.5 4.7 32.9 22.8 23.9
ergometrinine 0.9902 y = 3.6036x−12.6925 58.3 2.6 4.8 28.3 31.5 26.4
ergosine 0.9999 y = 2.1023x−9.7773 81.5 1.7 4.1 20.8 9.7 10.2
ergosinine 0.9995 y = 2.6077−16.4461 97.0 2.4 4.2 31.3 15.7 10.9
ergocornine 0.9996 y = 3.3136x−14.6219 67.7 2.2 4.6 24.6 12.3 14.1
ergocorninine 0.9996 y = 4.2199x−30.8679 99.9 0.8 1.8 24.4 5.7 11.0

Concentrations (μg kg−1) of ergot alkaloids determined in 16 positive compound feeds

Sample No. Err Errine Ert Ertine Erp Erpine Erm Ermine Ers Ersine Erc Ercine ∑ EAs
1 < LOQ     < LOQ < LOQ     23.2         23.2
2         < LOQ     22.2       < LOQ 22.2
3 < LOQ     < LOQ < LOQ   15.5         < LOQ 15.5
4             16.4 15.7         32.1
5   < LOQ                 24.4   24.4
6           < LOQ           < LOQ -
7   < LOQ                 < LOQ   -
8   < LOQ                 < LOQ   -
9         < LOQ < LOQ           < LOQ -
10         31.0     27.4         58.4
11         < LOQ     18.3 10.4       28.7
12               38.3   24.0     62.3
13     < LOQ < LOQ < LOQ < LOQ             -
14         < LOQ       < LOQ < LOQ < LOQ   -
15 < LOQ < LOQ     < LOQ     11.9         11.9
16         < LOQ 16.7             16.7

Validation results for recovery, repeatability and within-laboratory reproducibility

  Recovery (%) (at concentration:) Repeatability (%) (at concentration:) Reproducibility (%) (at concentration:)
  10 μg kg−1 250 μg kg−1 1,000 μg kg 1 10 μg kg−1 250 μg kg−1 1,000 μg kg 1 10 μg kg−1 250 μg kg−1 1,000 μg kg 1
ergocristine 102.8 94.5 95.8 3.7 7.9 9.2 8.1 5.4 7.3
ergocristinine 110.8 99.5 102.5 4.3 2.5 2.9 6.1 2.2 2.7
ergotamine 114.0 92.6 99.7 6.3 2.3 4.8 6.2 3.3 5.8
ergotaminine 117.8 96.2 102.4 1.2 3.9 1.4 3.4 4.2 2.5
ergocryptine 104.0 98.0 97.5 2.6 5.5 8.9 9.7 4.8 6.0
ergocryptinine 112.0 100.7 104.0 3.6 2.5 2.3 5.4 2.3 3.1
ergometrine 115.8 89.9 110.8 4.1 6.0 4.0 4.6 5.3 5.1
ergometrinine 111.9 85.2 112.6 3.3 4.3 4.7 7.7 5.5 4.0
ergosine 107.1 96.8 102.8 3.5 3.7 5.3 7.6 3.6 4.2
ergosinine 113.8 93.2 100.9 3.7 4.8 6.4 7.1 3.8 5.4
ergocornine 101.5 94.9 95.6 4.2 1.8 7.2 12.4 3.7 5.9
ergocorninine 110.2 99.6 104.5 3.9 3.2 1.9 6.7 2.8 3.1
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