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Peaks | tR (min) | Conc. (ng/mL) | Precursor Ion | Product Ion | |
---|---|---|---|---|---|
1. | Ascomycin | 1.45 | 10 | 809.53 | 756.50 |
2. | Tacrolimus | 1.47 | 10 | 821.61 | 768.51 |
3. | Sirolimus | 1.64 | 10 | 931.63 | 864.57 |
4. | Everolimus | 1.69 | 10 | 975.68 | 908.62 |
5. | Cyclosporin A | 1.83 | 10 | 1219.83 | 1202.87 |
6. | Cyclosporin D | 1.89 | 100 | 1233.91 | 1216.88 |
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.: | 70 °C | ||||||||||||||||||||
Inj. Vol.: | 5 µL | ||||||||||||||||||||
Mobile Phase | |||||||||||||||||||||
A: | 0.05% Formic acid, 5 mM ammonium formate in water | ||||||||||||||||||||
B: | Methanol | ||||||||||||||||||||
| |||||||||||||||||||||
Max Pressure: | 305 bar |
Detector | MS/MS |
---|---|
Ion Mode: | ESI+ |
Mode: | MRM |
Instrument | UHPLC |
Sample Preparation | Human whole blood was fortified with 4 immunosuppressive drugs at 10 ng/mL. For quantitation, cyclosporin D was used as the internal standard for cyclosporin A and ascomycin was used as the internal standard for tacrolimus, sirolimus, and everolimus. The blood sample (100 μL) was mixed with 200 μL of precipitation solution (1:4 v/v 0.2 M ZnSO4:methanol) containing 50 ng/mL of cyclosporin D and 5 ng/mL of ascomycin. The mixture was vortexed for 20 seconds at 3000 rpm and then centrifuged for 10 minutes at 4300 rpm. The supernatant was directly injected (5 μL) for analysis. |
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