2.5. Salivary Cortisol and Cortisone
Salivary samples were obtained using a Salivette kit (Sarstedt, Nümbrecht, Germany) to measure cortisol and cortisone concentrations. Once collected, samples were stored at 4 °C before being analyzed. Salivary cortisol and cortisone were measured by LC-MS/MS (Prominence liquid chromatography system; Shimadzu, Nakagyo, Japan; and 5500 Qtrap detector, Sciex, Framingham, MA, USA). A liquefying agent (Sputasol; Thermo Fisher Scientific, Waltham, MA, USA) was added to 400 µL of saliva sample, which was then incubated for 30 min at 37 °C. Next, solid-phase extraction with a hydrophilic lipophilic balance (Waters