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Pyrrolizidine alkaloids in honey: determination with liquid chromatography-mass spectrometry method

Ewelina Kowalczyk
Ewelina Kowalczyk
 and 
Krzysztof Kwiatek
Krzysztof Kwiatek
   | Jul 07, 2018
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Published Online: Jul 07, 2018
Page range: 173 - 181
Received: Feb 08, 2018
Accepted: Jun 20, 2018
DOI: https://doi.org/10.2478/jvetres-2018-0027
Keywords
© 2018 Ewelina Kowalczyk et al., published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

Fig. 1

SIM chromatograms of honey samples obtained for a) HF Bond cartridge and 3% ammonia in methanol elution (PAs concentration 1 μg kg−1); b) MCX cartridge and new elution mixture consisting of ethyl acetate, methanol, acetonitrile, ammonia, and triethylamine (8:1:1:0.1:0.1, v/v) elution (PAs concentration 1 μg kg−1); c) blank sampleInt – Intermedine, Lyc – lycopsamine, Jcb – jacobine, Ret – retrorsine, Hel – heliotrine, Snc – seneciphylline, Sen – senecionine, Ech – echimidine, Skn – senkirkine, Las – lasiocarpine
SIM chromatograms of honey samples obtained for a) HF Bond cartridge and 3% ammonia in methanol elution (PAs concentration 1 μg kg−1); b) MCX cartridge and new elution mixture consisting of ethyl acetate, methanol, acetonitrile, ammonia, and triethylamine (8:1:1:0.1:0.1, v/v) elution (PAs concentration 1 μg kg−1); c) blank sampleInt – Intermedine, Lyc – lycopsamine, Jcb – jacobine, Ret – retrorsine, Hel – heliotrine, Snc – seneciphylline, Sen – senecionine, Ech – echimidine, Skn – senkirkine, Las – lasiocarpine

Fig. 2

Stability results of the target PAs in dry SPE-purified extracts of honey matrix, stored at −18°C for 30 days
Stability results of the target PAs in dry SPE-purified extracts of honey matrix, stored at −18°C for 30 days

Fig. 3

The content of pyrrolizidine alkaloids in positive honey samples
The content of pyrrolizidine alkaloids in positive honey samples

Fig. 4

Recoveries of the alkaloids from different SPE cartridges eluted with the new solvents mixture consisting of ethyl acetate, methanol, acetonitrile, ammonia, and triethylamine (8:1:1:0.1:0.1, v/v)
Recoveries of the alkaloids from different SPE cartridges eluted with the new solvents mixture consisting of ethyl acetate, methanol, acetonitrile, ammonia, and triethylamine (8:1:1:0.1:0.1, v/v)

Selected (m/z) ions monitored and retention times of the monitored alkaloids. Coefficients of determination obtained for matrix calibration curves in a concentration range corresponding to 0–50 μg kg-–1 and matrix effect results

PAm/zRTR2ME (%)
Intermedine300.16.160.997105.5
Lycopsamine300.06.380.99116.5
Jacobine352.16.910.998119.2
Retrorsine352.18.310.998117.4
Heliotrine314.18.630.998119.9
Seneciphylline334.19.330.998119.2
Senecionine336.111.140.994122.6
Echimidine398.013.180.998117.1
Senkirkine366.113.370.995106.2
Lasiocarpine412.115.110.996107.5

Validation parameters evaluated for ten alkaloids in honey matrix

Concentration level (μg kg–1)Concentration level (μg kg–1)Concentration level (μg kg–1)
110501105011050
PARecovery (%)Repeatability RSD (%)Reproducibility RSD (%)
Intermedine80.684.696.314.66.05.417.716.610.1
Lycopsamine108.297.4100.611.19.12.516.39.27.1
Jacobine90.799.397.510.99.74.515.211.54.9
Retrorsine98.2105.598.613.65.27.516.87.38.5
Heliotrine80.9101.495.68.54.04.315.07.35.1
Seneciphylline108.1101.5111.67.35.32.512.811.215.6
Senecionine87.2106.9102.47.57.22.310.311.27.6
Echimidine103.9109.4114.58.15.66.615.217.013.0
Senkirkine88.0104.5100.59.12.94.411.89.55.3
Lasiocarpine82.2108.9105.62.912.43.112.112.57.9

Summary of detected PAs

PANumber of samplesMean concentration (μg kg–1)Median (μg kg–1)Min.-Max content (μg kg–1)
Intermedine44.81.9<LOQ–14.7
Lycopsamine59.43.01.7–32.9
Retrorsine41.11.2< LOQ–1.8
Senecionine31.0<LOQ< LOQ–2.9
Echimidine73.33.81.4–-5.2

Combinations of solvents used during optimisation of solid phase extraction step. Recovery rates obtained for target alkaloids at the concentration of 20 μg kg–1

SolventsRatio (v/v)Recovery (%)
EtAc : MeOH : NH4OH8 : 2 : 0.277–93
EtAc : MeOH : NH4OH : TEA8 : 2 : 0.1 : 0.184–98
EtAc : MeOH : ACN : NH4OH : TEA8: 1 : 1 : 0.1 : 0.179–105
EtAc : MeOH : ACN : NH4OH : TEA7: 1.5 : 1.5 : 0.1 : 0.178–92
EtAc : MeOH : ACN : NH4OH : TEA8 : 1 : 1 : 0.2 : 0.176–97
EtAc : MeOH : ACN : NH4OH : TEA3 : 3 : 3 : 0.1 : 0.161–93
MeOH : NH4OH11 : 1.548–110
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