Investigation on possible transformations of cortisol, cortisone and cortisol glucuronide in bovine faecal matter using liquid chromatography–mass spectrometry

Given the close resemblance of the ring A structure of prednisolone and prednisone on the one hand, and of androstadienedione on the other, the transformation of cortisol and cortisone into prednisolone and prednisone in cattle faeces was evaluated. A simple method that does not involve extraction but only the 1:100 dilution of cattle faeces, spiking with 400 ng/mL cortisol, cortisone or cortisol glucuronide and incubation of the suspension, was used. The analyses were performed by HPLC–MS3 to detect the supposed Δ1 dehydrogenation of the glucocorticoids. The decision limits (CCα) and detection capabilities (CCβ) were 2.0 and 3.0 ng/mL for cortisol, cortisone and prednisolone, 3.0 and 4.0 ng/mL for cortisol glucuronide and 7.0 and 10.0 ng/mL for prednisone, respectively. Intra-day and inter-day coefficients of variation (CV%), were 5.6–6.2 and 5.2–6.6 for cortisol glucuronide, cortisol, cortisone and prednisolone, and 16.0 and 16.2 for prednisone, respectively. The recoveries were in the range 110–143% for all analytes. Regression coefficients (R2) were in the range 0.996–0.999 for all analytes. The results show the hydrolysis of the conjugated form and the dehydrogenation in ring A in diluted faeces. It is therefore predicted that urine contaminated with faeces may be positive for prednisone and prednisolone in the same way as they are positive for boldenone, i.e. as a result of microbiological dehydrogenase activity on cortisol and cortisone.