Simplified Liquid Chromatography-Mass Spectrometry Methods for Gestagen Analysis in Animal Fat and Liver

Gestagens, a classof veterinary drugs also called progestogens,are synthetic hormones used to increase feed efficiency and rate ofgain in heifers. The Canadian Food Inspection Agency analyzes progestogensmelengestrol acetate (MGA), megestrol acetate, and chlormadinone acetateusing liquid chromatography-mass spectrometry (LC-MS).Our conventional gestagen method for kidney fat has many time-consumingsteps, including solid-phase extraction. A sample preparation procedurehaving fewer clean-up steps was developed for routine diagnostic analysisof kidney fat and provided similar results faster, and at lower cost.A confirmatory liver method for gestagens, developed using salt-assistedextraction, employed minimal clean-up steps that resulted in highchemical background at the desired lower limit of quantification (LLOQ).Differential ion mobility spectrometry, specifically high-field asymmetricwaveform ion mobility spectrometry (FAIMS), was used to filter chemicalbackground in the gas phase. The effect of the ionization probe positionon FAIMS parameters, including sensitivity, is described. With LC-FAIMS-MS,chemical background for each gestagen was virtually eliminated, resultingin a quantitative liver method having the desired 0.6 ng/g LLOQ andestimated limits of detection (LODs) up to 140 times lower than LC-MS.Incurred MGA samples, analyzed using kidney fat and liver methodsfrom the same animal, show levels within the quantitative ranges ofboth methods.