Determination of ent-kaurene in subcutaneous fat of Iberian pigs by gas chromatography multi-stage mass spectrometry with the aim to differentiate between intensive and extensive fattening systems

This work presents a gas chromatography multi-stage mass spectrometry (GC–MS3) method for the determination of ent-kaurene in subcutaneous fat of Iberian pig, present in adipose tissue of animals due to pasture ingestion (extensive fattening system). The method comprises a saponification and a liquid–liquid extraction of the unsaponifiable fraction, followed by an isolation of the hydrocarbon fraction by high performance liquid chromatography (HPLC) and analysis by GC–MS3 (ion trap) with electronic ionization. The GC–MS3 analysis allows the isolation and characterization of specific fragments from the original (MS1) molecular structure, and particularly, those fragments originated from the precursor ion (m/z = 229) characteristic of ent-kaurene. The MS/MS product fragment m/z = 213 is used as a further precursor fragment giving rise to a MS3 spectrum specific for ent-kaurene. The limit of detection of the MS3 technique is lower than 0.2 μg kg−1 and a linear regression has been found between 0.2 and 112 μg kg−1. This method is applicable for the determination of the fattening system of the Iberian pig.