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EtG/EtS on Raptor EtG/EtS: Updated Method Conditions (Mobile Phase and Gradient)

PeakstR (min)Conc.
Precursor IonProduct IonProduct Ion
1.Ethyl-β-D-glucuronide-d5 (EtG-d5)1.21200226.285.0-
2.Ethyl-β-D-glucuronide (EtG)1.23500221.275.185.1
3.Ethyl sulfate-d5 (EtS-d5)2.3650130.198.0-
4.Ethyl sulfate (EtS)2.38500125.197.180.0
ColumnRaptor EtG/EtS (cat.# 9325A12)
Dimensions:100 mm x 2.1 mm ID
Particle Size:2.7 µm
Pore Size:90 Å
Guard Column:UltraShield UHPLC precolumn filter, 0.2 μm frit (cat.# 25809)
Temp.:35 °C
Standard/SampleEthyl-β-D-glucuronide-d5 (EtG-d5) (cat.# 34102)
Ethyl-β-D-glucuronide (EtG) (cat.# 34101)
Ethyl sulfate-d5 sodium salt (EtS-d5) (cat.# 34104)
Ethyl sulfate sodium salt (EtS) (cat.# 34103)
Diluent:0.01% Formic acid in water
Conc.:500 ng/mL (40x acetonitrile precipitation sample prep)
Inj. Vol.:10 µL
Mobile Phase
A:0.01% Formic acid in water
B:0.1% Formic acid in acetonitrile
Time (min)Flow (mL/min)%A%B
Ion Source:Electrospray
Ion Mode:ESI-
Sample PreparationA 500 ng/mL standard was prepared in urine. A 50 μL aliquot was mixed with 10 μL of internal standard (20 μg/mL EtG-d5 and 5 μg/mL EtS-d5 in water) and 150 µL of acetonitrile by vortexing at 3000 rpm for 10 seconds and centrifuged at 4300 rpm for 10 minutes at 10 °C. After centrifugation, 100 µL of the supernatant was diluted with 900 µL (40x dilution) of 0.01% formic acid in water. The sample was then vortexed at 3000 rpm for 10 seconds and injected for LC-MS/MS analysis.