Determination of cinnamaldehyde, carvacrol and thymol in feedstuff additives by pressurized liquid extraction followed by gas chromatography-mass spectrometry

Specific blends of essential oils (BEOs) are promising substitutes for antibiotics to promote livestock performance and to reduce the incidence of intestinal disorders. Microencapsulation of BEOs has shown to improve their stability, bioavailability and to control their release rate once they are added to the feedstuff. The development and validation of a method for determining essential oil components such as carvacrol, thymol and cinnamaldehyde in a microencapsulated material used as feed additive is presented. Analytes were extracted from feed additives and feedstuff by pressurized liquid extraction (PLE) with methanol at 50 °C for 5 min. Methanol provided good recovery values and cleaner extracts than other polar organic solvents tested. However, for certain kind of composite additives ethyl acetate showed to be a better option because trans-cinnamaldehyde undergoes chemical reaction in methanol. Then PLE extracts were analysed by gas chromatography coupled to ion trap mass spectrometry in selected ion storage (SIS) mode. The analyte stability and the absence of analyte losses during the PLE process was checked by a recovery study. Also, the matrix effect was studied to assess accuracy. Recovery values were between 85 and 115% in most cases. Intra- and inter-day relative standard deviation values were less than 4 and 14%, respectively. Finally, the developed method was applied to the analysis of a microencapsulated feed additive, several composite feed additive samples containing microencapsulated BEOs and a spiked feedstuff, for quality control and in stability studies.