Analysis of steroid acids in crude oils by mixed-mode solid phase extraction coupled to gas chromatography-mass spectrometry

High molecular weight carboxylic acids (HMw-CAs), including steroid acids and hopanoic acids, are unique biomarker compounds in petroleum samples. This study introduces a mixed-mode solid phase extraction (SPE) method for the separation of these acids from crude oils. By combining hydrophobic and ion-exchange interactions, the mixed-mode SPE achieved superior recovery for separation of HMw-CAs compared to the conventional ion-exchange sorbent. The optimized SPE was coupled with gas chromatography-mass spectrometry to characterize individual steroid acids in biodegraded oils. As a result, 7 steroid acids with carboxyl groups on C-3 position were identified. Quantitative assessments of steroid acid concentrations across samples with varying biodegradation levels indicate that these acids likely originated during early diagenesis rather than forming through in-reservoir biodegradation of steranes. Notably, the observed resistance of these steroid acids to biodegradation suggests their potential as molecular markers for oil-source correlation in severely biodegraded petroleum systems.