Pyrrolizidine alkaloids (PAs) are probably the most widespread toxins of natural origin. More than 6,000 plant species produce these toxic compounds. Bees can forage on flowers of plants producing PAs, which leads to contamination of honey with the toxic compounds. To determine the contamination of honey with PAs, a sensitive method based on liquid chromatography coupled with mass spectrometry has been developed. PAs were extracted with 0.05 M sulphuric acid and purified with MCX cartridges. A solvent mixture consisting of ethyl acetate, methanol, acetonitrile, ammonia, and triethylamine (8:1:1:0.1:0.1, v/v) was used to wash alkaloids from the cartridges. After evaporation the residues were reconstituted in water and methanol mixture and subjected to LC–MS analysis. The developed method was validated according to SANTE/11945/2015 requirements. The recovery was from 80.6% to 114.5%. The repeatability ranged from 2.3% to 14.6%, and the reproducibility was from 4.9% to 17.7%. A new method for the determination of PAs in honey has been developed and validated. All evaluated parameters were in accordance with the SANTE/11945/2015 guidance document. Out of 50 analysed honey samples, 16 (32%) were positive for the content of at least one PA.

Pyrrolizidine alkaloids (PAs) are probably the most widespread toxins of natural origin. More than 6,000 plant species produce these toxic compounds. Bees can forage on flowers of plants producing PAs, which leads to contamination of honey with the toxic compounds. To determine the contamination of honey with PAs, a sensitive method based on liquid chromatography coupled with mass spectrometry has been developed.

Introduction: The main problem in determination of chloramphenicol in food of animal origin is a large number of matrices. The main target of this study was to create a method for determination and confirmation of chloramphenicol in products and food of animal origin. Material and Methods: Each 5 g matrix sample was mixed with 5 mL of water and 10 mL of acetonitrile/ethyl acetate, homogenised, and centrifuged. The organic layer was evaporated and redissolved in 6 mL of 4% NaCl. The extract was cleaned up by SPE technique. Chloramphenicol was analysed by LC-MS/MS in electrospray mode. Results: The procedure was validated according to the Commission Decision No. 2002/657/EC. The apparent recoveries were in the range of 92.1% to 107.1% with a repeatability less than 11.0% (4.4%-11.0%) and within-laboratory reproducibility below 13.6% (4.7%-13.6%). Conclusion: The method was successfully validated and proved to be efficient, precise, and useful for quantification of chloramphenicol in more than 20 different matrices.

Introduction: The main problem in determination of chloramphenicol in food of animal origin is a large number of matrices. The main target of this study was to create a method for determination and confirmation of chloramphenicol in products and food of animal origin. Material and Methods: Each 5 g matrix sample was mixed with 5 mL of water and 10 mL of acetonitrile/ethyl acetate, homogenised, and centrifuged. The organic layer was evaporated and redissolved in 6 mL of 4% NaCl. The extract was cleaned up by SPE technique. Chloramphenicol was analysed by LC-MS/MS in electrospray mode. Results: The procedure was validated according to the Commission Decision No. 2002/657/EC. The apparent recoveries were in the range of 92.1% to 107.1% with a repeatability less than 11.0% (4.4%-11.0%) and within-laboratory reproducibility below 13.6% (4.7%-13.6%). Conclusion: The method was successfully validated and proved to be efficient, precise, and useful for quantification of chloramphenicol in more than 20 different matrices.

Introduction: In the European Union the use of steroid growth promoters is prohibited under Council Directive 96/22/EC. For effective control of illegal use of natural steroids, highly sensitive analytical methods are required, because sex hormones can be present in very low concentrations in biological samples. The aim of the study was to develop a confirmatory method for the detection of testosterone in bovine serum at ppt level. Material and Methods: 17β-testosterone and internal standards of 17β-testosterone-d2 were extracted from serum samples with a mixture of tert-butyl methyl ether/petroleum ether and were directly analysed by an LC/MS/MS on QTRAP 5500 instrument with a TurboIon-Spray source operating in a positive ionisation mode. Chromatographic separation was achieved on the analytical column Inertsil® ODS-3 with an isocratic elution using mobile phase consisting of acetonitrile, methanol, and water. Method validation has been carried out in accordance with the Commission Decision 2002/657/EC. Results: The method was characterised by good recovery (82%) and precision (R.S.D 17 %). Decision limit (CCα) and detection capability (CCβ) was 0.05 μg L⁻¹ and 0.09 μg L⁻¹ respectively. The method met the criteria set out in Commission Decision 2002/657/EC for the purpose of confirmation in terms of retention time and ion ratio in the whole range of its application. Conclusions: The developed method is specific and sensitive, suitable for measuring the natural level of testosterone in blood of cattle and for use in routine control programme for the detection of this hormone in bovine serum.

Introduction: In the European Union the use of steroid growth promoters is prohibited under Council Directive 96/22/EC. For effective control of illegal use of natural steroids, highly sensitive analytical methods are required, because sex hormones can be present in very low concentrations in biological samples. The aim of the study was to develop a confirmatory method for the detection of testosterone in bovine serum at ppt level.

This paper reports polychlorinated dibenzo-p-dioxin (PCDD), polychlorinated dibenzofuran (PCDF), and polychlorinated biphenyl (PCB) concentrations in fish collected from Polish and Vietnamese farms and the related risk for consumers. Altogether, 160 samples were analysed using an isotope dilution technique with high-resolution gas chromatography coupled with high-resolution mass spectrometry (HRGC-HRMS). To characterise the potential health risk associated with PCDD/F and dioxin-like polychlorinated biphenyl (DL-PCB) intake, doses ingested in two 100 g portions of fish by adults and children were calculated and expressed as the percentage of the tolerable weekly intake (TWI) newly established by the EFSA in November 2018 at 2 pg WHO-TEQ kg⁻¹ b.w. Generally, levels in fish muscles were low in relation to maximum limits (4), being in the range of 0.02–3.98 pg WHO-TEQ g⁻¹ wet weight (w.w.) for PCDD/F/DL-PCBs and 0.05–24.94 ng g⁻¹ w.w. for NDL-PCBs. The highest concentration was found in eel muscles. The least polluted were pangas and zanders and the levels were at the limits of quantification. Consumption of two portions of fish per week results in intakes of 9– 866% TWI by children and 4–286% TWI by adults. Frequent consumption of some species (for example eel and bream) can pose a health risk to vulnerable consumers and especially children and pregnant women.

This paper reports polychlorinated dibenzo-p-dioxin (PCDD), polychlorinated dibenzofuran (PCDF), and polychlorinated biphenyl (PCB) concentrations in fish collected from Polish and Vietnamese farms and the related risk for consumers.

Some azo dyes, including Sudans I–IV and Para Red, are genotoxic and may be biotransformed to cancerogenic aromatic amines. They are banned as food and feed additives, but their presence has been detected in food. Aromatic amines are also considered potentially toxic. Online EC–MS is a promising tool to study the transformation mechanisms of xenobiotics such as azo dyes. The aim of the study was to investigate emulation of how azo dyes are enzymatically transformed to amines with EC–MS. The reduction reactions of five azo dyes (Sudans I–IV and Para Red) were conducted using a glassy carbon working electrode and 0.1% formic acid in acetonitrile. Reduction results were compared with the literature and in silico to select preliminary candidates for metabolites. The LC-MS/MS method was used to confirm results obtained by electrochemical reactor. A limited number of pre-selected compounds were confirmed as azo dyes metabolites – aniline for Sudan I, aniline and 4-aminoazobenzene for Sudan III, o-toluidine for Sudan IV, and 4-nitroaniline for Para Red. No metabolites were found for Sudan II. Electrochemistry–mass spectrometry was successfully applied to azo dyes. This approach may be used to mimic the metabolism of azo dyes, and therefore predict products of biotransformation.

Some azo dyes, including Sudans I–IV and Para Red, are genotoxic and may be biotransformed to cancerogenic aromatic amines. They are banned as food and feed additives, but their presence has been detected in food. Aromatic amines are also considered potentially toxic. Online EC–MS is a promising tool to study the transformation mechanisms of xenobiotics such as azo dyes. The aim of the study was to investigate emulation of how azo dyes are enzymatically transformed to amines with EC–MS.

Introduction: The differentially expressed proteins between healthy cows and those with footrot were identified to explore changes in protein profiles associated with the disease. Material and Methods: Out of 36 cows selected for the experiment, 18 footrot-affected cows were included in the treatment group (group T) and 18 unaffected cows were included in the control group (group C). Plasma samples from groups T and C were subjected to two-dimensional electrophoresis analysis and differentially expressed proteins were identified by matrix-assisted laser desorption/ionisation tandem time-of-flight mass spectrometry. Bioinformatics, including gene ontology analysis and pathway analysis, was used for analysing all proteins. Results: Out of 63 spots identified by 2DE, 33 were selected for mass spectrum analysis, which identified 11 differentially expressed proteins in 26 spots. Footrot led to changes in profiles in plasma proteins that were classified to the pathway of inflammatory response, complement, and blood coagulation, among others. Conclusion: This study provides evidence of the defence mechanisms of cows with footrot to explore strategies for treatment.

Introduction: The differentially expressed proteins between healthy cows and those with footrot were identified to explore changes in protein profiles associated with the disease. Material and Methods: Out of 36 cows selected for the experiment, 18 footrot-affected cows were included in the treatment group (group T) and 18 unaffected cows were included in the control group (group C). Plasma samples from groups T and C were subjected to two-dimensional electrophoresis analysis and differentially expressed proteins were identified by matrix-assisted laser desorption/ionisation tandem time-of-flight mass spectrometry. Bioinformatics, including gene ontology analysis and pathway analysis, was used for analysing all proteins. Results: Out of 63 spots identified by 2DE, 33 were selected for mass spectrum analysis, which identified 11 differentially expressed proteins in 26 spots. Footrot led to changes in profiles in plasma proteins that were classified to the pathway of inflammatory response, complement, and blood coagulation, among others. Conclusion: This study provides evidence of the defence mechanisms of cows with footrot to explore strategies for treatment.

Introduction: The article presents a rapid and simple analytical procedure for determination of four tetracyclines (oxytetracycline, tetracycline, chlortetracycline, and doxycycline) in animal medicated feedingstuffs. Material and Methods: Two-gramme samples were extracted by a Na₂EDTA-McIlvaine buffer (pH 4)/methanol mixtures (40/60, v/v). The determination was achieved by liquid chromatography using a Zorbax Eclipse XDB C18 analytical column with mass spectrometer detection (LC-MS). Results: Recoveries of the antibiotics from spiked feed samples ranged from 78.2% to 113.5%. The LOD and LOQ for tetracyclines in feeds ranged from 2.8 to 4.2 and from 4.3 to 5.7 mg/kg, respectively. Conclusion: The method was successfully validated and proved to be efficient, precise, and useful for quantification of tetracyclines in medicated feedingstuffs.

Introduction: The article presents a rapid and simple analytical procedure for determination of four tetracyclines (oxytetracycline, tetracycline, chlortetracycline, and doxycycline) in animal medicated feedingstuffs. Material and Methods: Two-gramme samples were extracted by a Na2EDTA-McIlvaine buffer (pH 4)/methanol mixtures (40/60, v/v). The determination was achieved by liquid chromatography using a Zorbax Eclipse XDB C18 analytical column with mass spectrometer detection (LC-MS). Results: Recoveries of the antibiotics from spiked feed samples ranged from 78.2% to 113.5%. The LOD and LOQ for tetracyclines in feeds ranged from 2.8 to 4.2 and from 4.3 to 5.7 mg/kg, respectively. Conclusion: The method was successfully validated and proved to be efficient, precise, and useful for quantification of tetracyclines in medicated feedingstuffs.

Introduction: The objective of this study was to describe a pharmacokinetic–pharmacodynamic (PK/PD) approach for determination of a rational dosage of ampicillin (AMP) and depletion of the antibiotic residues in milk after intramammary administration to cows.Material and Methods: The cows came from different farms from the Lublin Province area. They (n = 9) received 5 g of the drug, containing 75 mg of AMP sodium in physiological solution, through a syringe tube by intramammary administration. Following single intramammary administration, the milk samples (5 mL) were collected after 2, 4, 6, 8, 10, 24, 36, 48, and 60 h. The liquid chromatography-mass spectrometry analysis was performed on the Agilent 1200 system connected to an AB Sciex API 4000™ mass spectrometer. The pharmacokinetic analysis of the concentrations of the antibiotic in milk was performed using software Phoenix® WinNonlin® 6.4. Calculations were made in non-compartmental (slopes, highest, amounts, and moments) and compartmental analysis.Results: The pharmacokinetic characteristics of AMP after intramammary administration indicate rapid elimination of the drug from milk. The mean residence time had a several-fold lower value than the designated elimination half-life and amounts to only 3.4 h. The concentration of the drug in the milk dropped relatively quickly and the process was very dynamic.Conclusion: The conducted research confirms the rationale of using the PK/PD model in order to verify the dosing regimen for other antibiotic groups and various indicators of the applied PK/PD model.

Introduction: The objective of this study was to describe a pharmacokinetic–pharmacodynamic (PK/PD) approach for determination of a rational dosage of ampicillin (AMP) and depletion of the antibiotic residues in milk after intramammary administration to cows.