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Peaks | tR (min) | Precursor Ion | Product Ion | Product Ion | |
---|---|---|---|---|---|
1. | Cortisol-d4 | 0.75 | 367.4 | 121.0 | - |
2. | Endogenous cortisol | 0.76 | 363.3 | 120.9 | 91.1 |
3. | Cortisone-d8 | 0.88 | 369.4 | 167.9 | - |
4. | Endogenous cortisone | 0.89 | 361.3 | 163.2 | 91.0 |
Column | Raptor Biphenyl (cat.# 9309A52) | ||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Dimensions: | 50 mm x 2.1 mm ID | ||||||||||||||||||||||||||||
Particle Size: | 2.7 µm | ||||||||||||||||||||||||||||
Pore Size: | 90 Å | ||||||||||||||||||||||||||||
Guard Column: | Raptor Biphenyl EXP guard column cartridge 5 mm, 2.1 mm ID, 2.7 µm (cat.# 9309A0252) | ||||||||||||||||||||||||||||
Temp.: | 40 °C | ||||||||||||||||||||||||||||
Standard/Sample | |||||||||||||||||||||||||||||
Diluent: | Mobile phase A | ||||||||||||||||||||||||||||
Conc.: | Calculated concentration is 105.1 ng/mL and 135.9 ng/mL for cortisol and cortisone, respectively | ||||||||||||||||||||||||||||
Inj. Vol.: | 10 µL | ||||||||||||||||||||||||||||
Mobile Phase | |||||||||||||||||||||||||||||
A: | Water, 0.1% formic acid | ||||||||||||||||||||||||||||
B: | Acetonitrile, 0.1% formic acid | ||||||||||||||||||||||||||||
| |||||||||||||||||||||||||||||
Max Pressure: | 400 bar |
Detector | MS/MS |
---|---|
Ion Mode: | ESI+ |
Mode: | MRM |
Instrument | UHPLC |
Sample Preparation | 1. Centrifuge female human urine for 5 min at 4500 rpm, 10 °C. 2. Aliquot 380 µL supernatant. Add 20 µL each of internal standard solution (1 µg/mL in methanol). 3. Load 200 μL of sample on to ISOLUTE SLE+ 200 µL supported liquid extraction plate (part# 820-0200-P01). 4. Apply a pulse of vacuum to initiate flow. 5. Wait 5 min for sample to completely absorb. 6. Extract samples with 1 mL of MTBE. Allow solvent to flow for 5 min under gravity. Apply vacuum for 10-30 sec to complete elution. 7. Evaporate extracts to dryness under a stream of nitrogen. 8. Reconstitute in 200 µL mobile phase A prior to analysis. 9. Vortex to mix. |
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