Determination of ten steroid hormones in animal waste manure and agricultural soil using inverse and integrated clean-up pressurized liquid extraction and gas chromatography-tandem mass spectrometry

This paper presents a novel analytical methodology for the simultaneous determination of nine native and one synthetic steroid hormone in animal manure and agricultural soil. The optimized method applies pressurized liquid extraction (PLE) on manure samples packed in a 22 mL PLE cell: firstly by flushing the sample with heptane to remove unwanted matrix components (inverse-PLE, i-PLE) and secondly, performing internal clean-up (ic-PLE) and eluting the steroid hormones by attaching an additional 11 mL PLE cell downstream packed with graphite, silica and alumina. The PLE extracts were further cleaned by a two step solid phase clean-up (aminopropyl and silica gel), and in combination with a one hour derivatization the samples could be analyzed utilizing a GC-MS/MS system with PTV injector. Cholesterol, sitosterol and coprostanols were added and monitored during method development experiments as surrogates for environmental interfering compounds. The optimization demonstrated that the surrogate interfering compounds could be fractionated from the steroid hormones during the i-PLE step. The absolute PLE recoveries for the steroid hormones were between 67 and 107% from 0.5 g manure spiked with 50 ng. The method was applied on several manure and agricultural soil samples, demonstrating high levels of steroid hormones in manure and amended soils.