Pyrrolizidine alkaloids (PAs) are secondary metabolites produced by many plant species. Due to their toxicity PAs can pose a risk to human and animal health. To detect the toxic compounds in feed materials a sensitive method based on liquid chromatography coupled with mass spectrometry has been developed. PAs were extracted with sulphuric acid and purified with cation exchange cartridges. A newly developed solvent mixture consisting of ethyl acetate, methanol, acetonitrile, ammonia, and triethylamine 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 guidelines. The recovery was from 84.1% to 112.9%, the repeatability ranged from 3.0% to 13.6%, and the reproducibility was from 4.8% to 18.9%. A sensitive and selective method for determination of PAs in feed materials has been developed and validated. All evaluated validation parameters were in accordance with EU Reference Laboratories document no. SANTE/11945/2015. Almost 41% of the analysed feed samples were positive for the presence 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. 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.

Objective-To evaluate the efficacy of an orally administered vaccine of Mycoplasma hyopneumoniae that was prepared by spray drying or solvent evaporation. Animals-Thirty 6-week-old, crossbred, specificpathogen- free (SPF) pigs. Procedure-Pigs were randomly allocated into 5 groups and housed in an SPF facility. Pigs in 2 groups (groups AQ and CAP) were fed M hyopneumoniae enteric-coated vaccine on days 0, 10, and 20. A third group (group IM) received an IM injection of M hyopneumoniae vaccine with aluminium hydroxide as an adjuvant on days 0, 10, and 20. The last 2 groups (nonvaccinated- challenged [NV-C] and nonchallenged [NC]) were fed a sham treatment. All 24 pigs in groups AQ, CAP, IM, and NV-C were challenge exposed with 5 ml of a 10% pneumonic lung suspension administered on day 40 via intubation of the trachea. All pigs were slaughtered and the lungs removed and examined for lesions on day 68. Results-In vitro studies indicated that these 2 microencapsulation techniques formed an effective shell and protected mycoplasmal antigen from gastric acid. Results of inoculation and challenge tests indicated that microencapsulated M hyopneumoniae were sufficiently potent to induce an immune response and provide good protection. Conclusions and Clinical Relevance-Orally administered microencapsulated M hyopneumoniae vaccines induced an immune response and reduced the severity of lung lesions in challengeexposed pigs. Results suggest that this novel method can be applied to other antigens, because the spray-drying process yielded an orally administered M hyopneumoniae vaccine that induced a good immune response.