5975c inert msd detector

Each derivatized sample (1 μL) was injected using the splitless mode with an Agilent Technologies 7890A chromatograph equipped with a HP-5MS column (30 m × 0.25 mm × 0.25 μm) and Agilent Technologies 5975C inert MSD detector. The initial oven temperature was held at 70 °C for 2 min, increased to 160 °C with 6 °C/min, and then to 240 °C with 10 °C/min, and finally increase to 300 °C with 20 °C/min, constant for 6 min, with He as carrier gas (1 mL/min) and MS detection. The temperatures of injector, transfer line, and electron impact ion source were set to 250, 290, and 230 °C, respectively.

Urinary free cortisol (F) and cortisone (E) quantification was performed using a validated liquid chromatography tandem mass spectrometry (LC-MS/MS) method, as extensively described elsewhere (21 (link)). Briefly, separation and quantification of both steroids was performed by using the MS urinary free cortisol/cortisone kit (ISBN-BSN, Castelleone, Italy) on a Nexera X2 series UHPLC (Shimadzu, Kyoto, Japan) coupled with a 4500 MD triple quadrupole MS (Sciex, Milan, Italy) detector. The mean intra- and inter-assay imprecision were between 1.7% and 11.3%
Total tetrahydro-cortisol (THF), 5α-tetrahydrocortisol (5α-THF), and tetrahydro-cortisone (THE) were quantified by gas chromatography–mass spectrometry (GC-MS) method, as previously described (17 (link), 22 (link)). Instrumentation used comprised an Agilent Technologies 7890A gas chromatograph and an Agilent Technologies 5975C inert MSD detector (Agilent Technologies Inc., Santa Clara, CA, USA). The mean intra- and inter-assay imprecision were between 3.9% and 13% for both methods.
Either F/E or THF+allo-THF/THE was measured on 24-h collected urine.

Dry leaf samples were subjected to alkaloids extraction as previously described previously (Goossens et al., 2003 ; Zhang et al., 2012 (link)) with minor modification. Briefly, 10mg of homogenized dry leaf sample was soaked in 1ml of 10% NaOH (w/v) for 20min and then extracted by vortexing with an equivalent volume of dichloromethane. The organic layer was collected after centrifugation. The nicotine content was measured on an Agilent Technologies 7890A Chromatograph equipped with a DB 5 MS column and Agilent Technologies 5975C inert MSD detector with helium as the carrier gas. The column temperature was held at 100 °C for 5min, increased to 210 °C at an increment of 50 °C min^–1, and then held at 210 °C for 4min. The ion source temperature was 230 °C and the quadrupole temperature was 150 °C. Nicotine from Sigma–Aldrich was used as the standard control.