LC/MS (TOF) Analysis of Drugs and Their Glucuronide Metabolites in Urine on Titan™ C18 after Solid Phase Extraction (SPE) using Supel™-Select SCX, β-Glucuronidase Enzyme Digestion

LC/MS (TOF) Analysis of Drugs and Their Glucuronide Metabolites in Urine on Titan™ C18 after Solid Phase Extraction (SPE) using Supel™-Select SCX, β-Glucuronidase Enzyme Digestion

Conditions

sample preparation Solid Phase Extraction (SPE)
sample/matrix human urine
extraction process urine samples were spiked at 200 ng/mL each analyte and digested with 10kU of beta-glucuronidase enzyme at 60 °C for 120 min
SPE tube/cartridge Supel-Select SCX, 60 mg/3 mL (54241-U)
condition 1 mL of 1% formic acid in acetonitrile followed by 1 mL water
sample addition 1.4 mLurine
washing 2 mL water followed by 1 mL of water:methanol (75:25)
elution 1 mL of 10% ammonium hydroxide in acetonitrile
eluate post-treatment evaporate to dryness and reconstitute in 1 mL water
column Titan C18, 5 cm x 2.1 mm I.D., 1.9 µm particles (577122-u)
column temp. 35 °C
mobile phase [A] 5 mM ammonium formate; [B] 5 mM ammonium formate in acetonitrile:water (95:5)
gradient 5 to 90% B in 2.5 min, held for 0.5 min, to 5% B in 0.1 min, held for 0.5 min
flow rate 0.3 mL/min
injection 2 µL
detector ESI(+) TOF, m/z 100-1000

Description

Analysis Note Sample pretreatment: urine samples were spiked at 200 ng/mL each analyte and digested with 10kU of beta-glucuronidase enzyme at 60 °C for 120 min, A set of compounds representing five drug classes that are routinely tested in urine were extracted from urine and analyzed using LC/MS (TOF). The entire workflow, from enzymatic digestion to hydrolyze the glucuronide metabolites to analysis using a Titan UHPLC column is presented here. Cerilliant CRMS provided reliable quantification.
Categories Analytical Chromatography, Clinical, Forensics and Toxicology, SPE Applications, steroids, Beta-blockers, Opiate, Anticonvulsants, anticonvulsants, Drugs of Abuse, Opiates, Drugs of abuse, Steroids, beta-blockers, analgesics, Analgesics, Anticonvulsant/antiepiletic/antiseizure, opiate, drugs of abuse
Featured Industry Clinical Research
Forensics and Toxicology
Legal Information Supel is a trademark of Sigma-Aldrich Co. LLC, Titan is a trademark of Sigma-Aldrich Co. LLC
suitability application for SPE, application for HPLC

Materials

     
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