Sample pretreatment

• Homogenize urine sample by stirring
• Fill 0.5 mL sample into a safe-lock tube
• Add standard solution, internal standard solution as described in table 1
• Add 25 µL acetic acid (25 %) and 1000 µL of a mixture of ethyl acetate – n-hexane (9+1, v+v)
• Shake vigorously for 30 sec
• Centrifuge at room temperature at 13400 rpm for 10 min
• Take up 500 µL of the organic phase for derivatization in a vial
• Concentrate organic phase at 40 °C under nitrogen stream to dryness
• Add 50 µL ethyl acetate and 50 µL MSTFA (REF 701270.201)
• Shake vigorously for 30 sec and incubate mixture at 80 °C for 30 min
• Take sample solution for GC-MS analysis

• Injection volume: 1 µL
• Injection mode: Splitless
• Injection temperature: 250 °C
• Carrier gas: Helium
• Flow: 1.31 mL/min
• Temperature: 70 °C [2 min] --> [20 °C/min] --> 250 °C [4 min] --> [20 °C/min] --> 300 °C [17 min]